Identification of coagulase-negative staphylococci isolated from caprine milk samples by PCR-RFLP of groEL gene

Onni, Toniangelo; Vidili, A.; Bandino, Ennio; Marogna, Gavino; Schianchi, S.; Tola, Sebastiana

doi:10.1016/j.smallrumres.2011.10.004

Cited by 24 publications

(19 citation statements)

References 29 publications

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“…To study differences in pathogenicity between staphylococcal species, accurate species identification methods are required. Several NAS species in goats are difficult to identify by phenotypic methods alone, yielding poor typeability; hence, genotypic methods are considered more reliable (Koop et al, 2012a;Onni et al, 2012). Sequencing of housekeeping genes or the whole genome is considered the reference method for identification of staphylococcal species (Zadoks and Watts, 2009).…”

Section: Use Of Maldi-tof To Characterize Staphylococcal Intramammarymentioning

confidence: 99%

“…Gene-based identification methods and other molecular-based techniques rely on the availability of a database, and typeability and accuracy are affected by the origin of the isolates used to build the database. Polymerase chain reaction RFLP analysis of the staphylococcal groEL gene (Onni et al, 2012), and transfer RNA-intergenic spacer PCR supplemented with gene sequence analysis were previously reported to have good accuracy and typeability for identification of NAS isolated from goat milk (McDougall et al, 2010;Koop et al, 2012b). Another rapid, cost-effective molecular method and potential alternative to gene sequencing is MALDI-TOF MS.…”

Section: Use Of Maldi-tof To Characterize Staphylococcal Intramammarymentioning

confidence: 99%

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Use of MALDI-TOF to characterize staphylococcal intramammary infections in dairy goats

Gosselin

Lovstad

Dufour

et al. 2018

Journal of Dairy Science

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The most common pathogens causing intramammary infections (IMI) in dairy goats are staphylococci. Gene sequencing has been the reference method for identification of staphylococcal species, but MALDI-TOF mass spectrometry could represent a rapid and cost-effective alternative method. The objectives were to evaluate the typeability and accuracy of partial gene sequencing and MALDI-TOF for identifying staphylococci isolated from caprine milk samples, and to evaluate the relationship between staphylococcal species IMI, milk somatic cell score (SCS), and milk yield (MY). A composite (goat-level) milk sample was collected from all 940 lactating goats in a single herd. Dairy Herd Information Association test-day data for parity, days in milk, SCS, and MY were retrieved from Dairy Herd Information Association records. Milk samples were cultured on Columbia blood agar, and isolates from samples that yielded a single colony type of a presumptively identified Staphylococcus spp. were identified by PCR amplification and partial sequencing of rpoB, tuf, or 16S-rRNA, and MALDI-TOF. Mixed linear models were used to evaluate the relationship between staphylococcal IMI, SCS, and MY. The goat-level prevalence of staphylococcal IMI based on isolation of a single colony type was 24.4% (213/874). Seventeen goats had a contaminated sample. Among the remaining goats (n = 857), the most common species causing single colony-type IMI were Staphylococcus simulans (7.9%), Staphylococcus xylosus (3.5%), Staphylococcus caprae (3.6%), Staphylococcus chromogenes (2.9%), and Staphylococcus epidermidis (2.2%). The typeability of staphylococcal isolates with partial housekeeping gene sequence analysis (rpoB, complemented by tuf and 16S as needed) was 97.7%. The typeability and accuracy of MALDI-TOF were 84 and 100%, respectively. Overall, only Staphylococcus chromogenes IMI was associated with a higher SCS than goats with no growth. After adjusting for parity and stage of lactation, staphylococcal IMI status was not significantly associated with MY. For the staphylococci isolated from goats in this herd, MALDI-TOF proved an accurate method of speciation with a relatively high typeability. An association between staphylococcal IMI, SCS, and MY was not defined using goat-level data with the exception of S. chromogenes IMI, which was associated with a higher SCS than goats with no growth.

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Section: Use Of Maldi-tof To Characterize Staphylococcal Intramammarymentioning

confidence: 99%

Section: Use Of Maldi-tof To Characterize Staphylococcal Intramammarymentioning

confidence: 99%

Use of MALDI-TOF to characterize staphylococcal intramammary infections in dairy goats

Gosselin

Lovstad

Dufour

et al. 2018

Journal of Dairy Science

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“…A major drawback for looking beyond the border of CNS as a group has long been the lack of consistent identification and typing methods for isolates from nonhuman sources. Indeed, the routine phenotypic methods that were traditionally used to identify CNS species from milk were essentially developed for use with human isolates and were proven unreliable in case of (some) CNS species found in milk (Thorberg and Brändström, 2000;Taponen et al, 2006Taponen et al, , 2008Capurro et al, 2009;Sampimon et al, 2009b;Onni et al, 2010Onni et al, , 2012Park et al, 2011a;Koop et al, 2012a). Recent studies have been able to overcome these difficulties as various validated molecular methods, including sequencing and fingerprinting methods, have become available (Piessens et al, 2010;Braem et al, 2011).…”

Section: Main Cns Found In Ruminant Milkmentioning

confidence: 99%

“…Therefore, the rest of this review will focus mainly on these species, which will further be referred to as the 5 main species. Sometimes other CNS species from milk will be considered in addition to the 5 main species, such as Staphylococcus caprae, being one of the main species in milk from goats and sheep (Onni et al, 2010(Onni et al, , 2012Koop et al, 2012a), as will CNS species from other niches than milk, including humans or the bovine environment. Studies with species data resulting from (solely) phenotypic identification will occasionally be cited.…”

Section: Main Cns Found In Ruminant Milkmentioning

confidence: 99%

Invited review: Effect, persistence, and virulence of coagulase-negative Staphylococcus species associated with ruminant udder health

Vanderhaeghen

Piepers

Leroy

et al. 2014

Journal of Dairy Science

155

149

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The aim of this review is to assess the effect of coagulase-negative staphylococci (CNS) species on udder health and milk yield in ruminants, and to evaluate the capacity of CNS to cause persistent intramammary infections (IMI). Furthermore, the literature on factors suspected of playing a role in the pathogenicity of IMI-associated CNS, such as biofilm formation and the presence of various putative virulence genes, is discussed. The focus is on the 5 CNS species that have been most frequently identified as causing bovine IMI using reliable molecular identification methods (Staphylococcus chromogenes, Staphylococcus simulans, Staphylococcus haemolyticus, Staphylococcus xylosus, and Staphylococcus epidermidis). Although the effect on somatic cell count and milk production is accepted to be generally limited or nonexistent for CNS as a group, indications are that the typical effects differ between CNS species and perhaps even strains. It has also become clear that many CNS species can cause persistent IMI, contrary to what has long been believed. However, this trait appears to be quite complicated, being partly strain dependent and partly dependent on the host's immunity. Consistent definitions of persistence and more uniform methods for testing this phenomenon will benefit future research. The factors explaining the anticipated differences in pathogenic behavior appear to be more difficult to evaluate. Biofilm formation and the presence of various staphylococcal virulence factors do not seem to (directly) influence the effect of CNS on IMI but the available information is indirect or insufficient to draw consistent conclusions. Future studies on the effect, persistence, and virulence of the different CNS species associated with IMI would benefit from using larger and perhaps even shared strain collections and from adjusting study designs to a common framework, as the large variation currently existing therein is a major problem. Also within-species variation should be investigated.

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“…Several molecular approaches have been suggested for the proper identification of CoNS, since phenotypic methods are timeconsuming and unreliable for animal samples (Park et al, 2011). PCR-RFLP of the groEL or hsp60 (heat-shock protein) gene has been effectively employed in the identification of CoNS from both bovine and caprine mastitis (Santos et al, 2008;Onni et al, 2012). Recently, the proteomic method MALDI-TOF MS has been increasingly used in microbial identification, due to its reliability, speed, and ease of use.…”

Section: Introductionmentioning

confidence: 99%

Characterization of Coagulase-Negative Staphylococci and pheno-genotypic beta lactam resistance evaluation in samples from bovine Intramammary infection

Melo

Motta

Rojas

et al. 2018

Arq. Bras. Med. Vet. Zootec.

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This study aimed to identify Coagulase-Negative Staphylococci (CoNS) species isolated from bovine mastitis, through phenotypic and PCR-RFLP (Restriction Fragment Length Polymorphism-Polimerase Chain Reaction) methods and to compare both techniques to matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) technique. Among them, the PCR-RFLP method, using a partially conserved sequence of the groEL gene, is a promising alternative, because of its reproducibility and reliability. On the other hand, the proteomic technique MALDI-TOF MS provides an accurate and much faster diagnosis and has been increasingly employed in microbiological identification. The pheno-genotypic profiles of beta-lactam resistance were also investigated, this characterization is important, considering that the use of antimicrobials is a key element for mastitis control in dairy farms. The concordance of the phenotypic, PCR-RFLP and MALDI-TOF MS assays to identify CoNS species was 77,5% (31/40). S. chromogenes was the species most frequently isolated. Antibiotic resistance rate was relatively low, registering values of 25.5% to penicillin, 9.6% to oxacillin and 6.2% to cefoxitin. Resistance to imipenem, cephalotin and amoxicillin+clavulanate was not observed. The mecA gene and its variant were detected in 7.6% and 4,1% of the isolates respectively. The blaZ gene was found in 43.2% of the strains resistant to penicillin.Keywords: Staphylococcus spp., bacterial resistance, PCR-RFLP and MALDI-TOF MS 77,5% (31/40 RESUMO Este estudo teve como objetivo identificar isolados de Staphylococcus coagulase-negativa (SCN) isolados de mastite bovina, por meio de métodos fenotípicos e PCR-RFLP (reação em cadeia de polimerase -polimorfismo nos fragmentos de restrição), e compará-los com a técnica de tempo de voo de ionização/desorção por laser assistida por matriz de espectrofotometria de massa (MALDI-TOF MS). O método de PCR-RFLP, que utiliza uma parte conservada da sequência do gene groEL, é uma alternativa promissora, por ser reprodutível e confiável. Por outro lado, a técnica proteômica MALDI-TOF MS permite uma acurácia e um diagnóstico muito mais rápidos e tem sido cada vez mais empregada na identificação microbiológica. Os perfis fenogenotípicos de resistência aos beta-lactâmicos também foram investigados. Essa caracterização é importante, considerando-se que os antimicrobianos são os elementos-chave para o controle da mastite na produção leiteira. A concordância entre os testes fenotípicos, PCR-RFLP E MALDI-TOF MS na identificação foi de

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Identification of coagulase-negative staphylococci isolated from caprine milk samples by PCR-RFLP of groEL gene

Cited by 24 publications

References 29 publications

Use of MALDI-TOF to characterize staphylococcal intramammary infections in dairy goats

Use of MALDI-TOF to characterize staphylococcal intramammary infections in dairy goats

Invited review: Effect, persistence, and virulence of coagulase-negative Staphylococcus species associated with ruminant udder health

Characterization of Coagulase-Negative Staphylococci and pheno-genotypic beta lactam resistance evaluation in samples from bovine Intramammary infection

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