Isolation of Phages and Study of their In vitro Efficacy on Staphylococcus aureus Isolates Originating from Bovine Subclinical Mastitis

2021 ◽  
Author(s):  
A.S. Srujana ◽  
J. Sonalika ◽  
D.S. Akhila ◽  
M.R. Juliet ◽  
P. Sheela
Keyword(s):  
Staphylococcus Aureus ◽  
Bovine Mastitis ◽  
Subclinical Mastitis ◽  
Global Market ◽  
Host Susceptibility ◽  
Health Concept ◽  
Disk Diffusion Assay ◽  
Potential Strategy ◽  
The One

Background: India leads the global market in milk production. However, bovine mastitis, which is the mammary gland inflammation in dairy cattle characterized by physical, chemical, bacteriological changes in milk results in commercial losses. Staphylococcus aureus, is the major causative agent. The treatment of mastitis caused by this pathogen is mainly by antibiotics. Emphasizing on the one health concept, phage therapy is an appropriate alternative to antibiotic. The present study was aimed to isolate and characterize bacteriophages against Staphylococcus aureus associated with bovine mastitis.Methods: Thirty two isolates of S. aureus obtained from the milk of mastitis cows were characterized by phenotypic and genotypic methods. Antibiotic susceptibility of the isolates was carried out by disk diffusion assay. Milk and cow shed wastewater were used for phage isolation. Phages were characterized by host susceptibility and RAPD assay.Result: Nineteen phages were isolated from the cowshed waste water. All the milk samples showed negative for the presence of phage. The phage 24 (A2) which had the broadest host range, was selected for the CFU drop assay. The phage was able to clear the lawn of S. aureus culture when grown on agar at different time points thus indicating that topical application of this phage would be a potential strategy to control S. aureus infection leading to mastitis. This study provides a basis to continue the exploration of the potential of PSA2 phage as a candidate for the treatment of Staphylococcal mastitis.

scholarly journals Within-Host Adaptation of Staphylococcus aureus in a Bovine Mastitis Infection Is Associated with Increased Cytotoxicity

2021 ◽  
Vol 22 (16) ◽  
pp. 8840
Author(s):  
Katharina Mayer ◽  
Martin Kucklick ◽  
Helene Marbach ◽  
Monika Ehling-Schulz ◽  
Susanne Engelmann ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Bovine Mastitis ◽  
Adaptive Strategy ◽  
Subclinical Mastitis ◽  
Growth Conditions ◽  
Typical Feature ◽  
Host Adaptation ◽  
Bovine Mammary Epithelial Cells ◽  
Strain Pair

Within-host adaptation is a typical feature of chronic, persistent Staphylococcus aureus infections. Research projects addressing adaptive changes due to bacterial in-host evolution increase our understanding of the pathogen’s strategies to survive and persist for a long time in various hosts such as human and bovine. In this study, we investigated the adaptive processes of S. aureus during chronic, persistent bovine mastitis using a previously isolated isogenic strain pair from a dairy cow with chronic, subclinical mastitis, in which the last variant (host-adapted, Sigma factor SigB-deficient) quickly replaced the initial, dominant variant. The strain pair was cultivated under specific in vitro infection-relevant growth-limiting conditions (iron-depleted RPMI under oxygen limitation). We used a combinatory approach of surfaceomics, molecular spectroscopic fingerprinting and in vitro phenotypic assays. Cellular cytotoxicity assays using red blood cells and bovine mammary epithelial cells (MAC-T) revealed changes towards a more cytotoxic phenotype in the host-adapted isolate with an increased alpha-hemolysin (α-toxin) secretion, suggesting an improved capacity to penetrate and disseminate the udder tissue. Our results foster the hypothesis that within-host evolved SigB-deficiency favours extracellular persistence in S. aureus infections. Here, we provide new insights into one possible adaptive strategy employed by S. aureus during chronic, bovine mastitis, and we emphasise the need to analyse genotype–phenotype associations under different infection-relevant growth conditions.

scholarly journals Microbiological and molecular aspects of Staphylococcus aureus isolated from bovine mastitis in West-Central Brazil

2020 ◽  
Vol 14 (1) ◽  
pp. 87
Author(s):  
T. B. Vieira ◽  
R. Almeida ◽  
I. B. Jesus ◽  
F. Freitas ◽  
R. T. Kemper ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Antimicrobial Agents ◽  
Bovine Mastitis ◽  
Subclinical Mastitis ◽  
Enterotoxin Gene ◽  
Central Brazil ◽  
Lactating Cows ◽  
Molecular Aspects ◽  
And Staphylococcus Aureus

Bovine mastitis is a major disease affecting dairy cattle, and Staphylococcus aureus is one of the most important agent involved in this condition due to its capacity to produce enterotoxins and develop resistance to antimicrobial agents. This study aimed to detect S. aureus strains in milk samples from cows with subclinical mastitis employing microbiological and molecular analysis.  Eleven farms were visited and from 187 lactating cows sampled, 33 S. aureus strains were isolated.  Only one of the 33 strains was positive for mecA resistance gene, 23 were positive for sea enterotoxin gene, and none was positive for seb or sec enterotoxin gene. S. aureus strains were submitted to in vitro antimicrobial susceptibility test and 63.6% (21/33) were susceptible to all antimicrobials tested, while 36.3% (12/33) were resistant to one or more antimicrobial agents. Identification of mecA and the sea genes highlighted the need to elaborate strategies to reduce problems related to animal. Furthermore, the identification of bovine mastitis caused by S. aureus is very important to manage herd and to public health, since milk contaminated by this pathogen can lead to serious health problems.

scholarly journals Expression of icaA and icaD genes in biofilm formation in Staphylococcus aureus isolates from bovine subclinical mastitis

2021 ◽  
Vol 41 ◽  
Author(s):  
Viviane F. Marques ◽  
Huarrisson A. Santos ◽  
Thomas H. Santos ◽  
Dayanne A. Melo ◽  
Shana M.O. Coelho ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Biofilm Formation ◽  
Phenotypic Expression ◽  
Bovine Mastitis ◽  
Subclinical Mastitis ◽  
Transcriptional Analysis ◽  
Important Species ◽  
High Prevalence ◽  
Staphylococcus Spp

ABSTRACT: Staphylococcus spp. plays a significant role in the etiology of bovine mastitis. Staphylococcus aureus is considered the most important species due to the high prevalence and the difficulty of in vivo treatment that is related to the expression of virulence factors and biofilm formation. This study aimed to detect the phenotypic expression of the biofilm formation in 20 S. aureus isolated from bovine mastitis and to evaluate the expression and regulation of genes involved in its production. MALDI-TOF and phenogenotypic identification assays were performed to characterize the isolates. The phenotypic biofilm production and the presence of icaA and icaD and bap genes were evaluated. The Agr system was typified (agr I, agr II, agr III and agr IV) and its regulator (agr RNAIII) was detected. Furtherly, Real-time PCR (qPCR) was performed at chosen times to quantify the expression of icaA, icaD and hld genes in three selected isolates. All 20 strains were biofilm producers and most presented icaA and icaD genes. Only one isolate presented the bap gene. The agr gene type II showed a prevalence of 70%. Transcriptional analysis revealed increased expression of ica genes at eight hours of growth. These results confirm that polysaccharides production mediated by the icaADBC operon genes is an essential mechanism to the biofilm formation and contributes to the early stages of bacterial growth.

scholarly journals Efficacy of Bacteriophages Against Staphylococcus aureus Isolates from Bovine Mastitis

2020 ◽  
Vol 13 (3) ◽  
pp. 35 ◽  
Author(s):  
Isabel Titze ◽  
Tatiana Lehnherr ◽  
Hansjörg Lehnherr ◽  
Volker Krömker
Keyword(s):  
Staphylococcus Aureus ◽  
Host Range ◽  
Bacterial Infections ◽  
Plaque Assay ◽  
Bovine Milk ◽  
Bovine Mastitis ◽  
Raw Milk ◽  
Pasteurized Milk

The lytic efficacy of bacteriophages against Staphylococcus aureus isolates from bovine milk was investigated in vitro, regarding possible applications in the therapy of udder inflammation caused by bacterial infections (mastitis). The host range of sequenced, lytic bacteriophages was determined against a collection of 92 Staphylococcus (S.) aureus isolates. The isolates originated from quarter foremilk samples of clinical and subclinical mastitis cases. A spot test and a subsequent plaque assay were used to determine the phage host range. According to their host range, propagation and storage properties, three phages, STA1.ST29, EB1.ST11, and EB1.ST27, were selected for preparing a bacteriophage mixture (1:1:1), which was examined for its lytic activity against S. aureus in pasteurized and raw milk. It was found that almost two thirds of the isolates could be lysed by at least one of the tested phages. The bacteriophage mixture was able to reduce the S. aureus germ density in pasteurized milk and its reduction ability was maintained in raw milk, with only a moderate decrease compared to the results in pasteurized milk. The significant reduction ability of the phage mixture in raw milk promotes further in vivo investigation.

scholarly journals Dalbavancin, Vancomycin and Daptomycin Alone and in Combination with Cefazolin against Resistant Phenotypes of Staphylococcus aureus in a Pharmacokinetic/Pharmacodynamic Model

Antibiotics ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 696 ◽  
Author(s):  
Jacinda C. Abdul-Mutakabbir ◽  
Razieh Kebriaei ◽  
Kyle C. Stamper ◽  
Zain Sheikh ◽  
Philip T. Maassen ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Bactericidal Activity ◽  
Inhibitory Concentration ◽  
Pharmacodynamic Model ◽  
In Vitro Tests ◽  
Beta Lactam ◽  
Fold Reduction ◽  
The One ◽  
Time Kill

The most efficacious antimicrobial therapy to aid in the successful elimination of resistant S. aureus infections is unknown. In this study, we evaluated varying phenotypes of S. aureus against dalbavancin (DAL), vancomycin (VAN), and daptomycin (DAP) alone and in combination with cefazolin (CFZ). The objective of this study was to observe whether there was a therapeutic improvement in adding a beta-lactam to a glycopeptide, lipopeptide, or a lipoglycopeptide. We completed a series of in vitro tests including minimum inhibitory concentration testing (MIC) of the antimicrobials in combination, time-kill analysis (TKA), and a 168 h (7-day) one-compartment pharmacokinetic/pharmacodynamic (PK/PD) model on two daptomycin non-susceptible (DNS), vancomycin intermediate S. aureus strains (VISA), D712 and 6913. Results from our MIC testing demonstrated a minimum 2-fold and a maximum 32-fold reduction in MIC values for DAL, VAN, and DAP in combination with CFZ, in contrast to either agent used alone. The TKAs completed on four strains paralleled the enhanced activity demonstrated via the combination MICs. In the one-compartment PK/PD models, the combination of DAP plus CFZ or VAN plus CFZ resulted in a significant (p < 0.001) improvement in bactericidal activity and overall reduction in CFU/ml over the 7-day period. While the addition of CFZ to DAL improved time to bactericidal activity, DAL alone demonstrated equal and more sustained overall activity compared to all other treatments. The use of DAL alone, with or without CFZ and the combinations of VAN or DAP with CFZ appear to result in increased bactericidal activity against various recalcitrant S. aureus phenotypes.

scholarly journals Pantothenate-requiring dwarf colony variants ofStaphylococcus aureusas the etiological agent in bovine mastitis

1969 ◽  
Vol 67 (3) ◽  
pp. 511-516 ◽  
Author(s):  
D. Sompolinsky ◽  
I. Gluskin ◽  
G. Ziv
Keyword(s):  
Staphylococcus Aureus ◽  
Metabolic Disorder ◽  
Bovine Mastitis ◽  
Etiological Agent

SUMMARYIn a herd affected with bovine mastitis, dwarf colony variants ofStaphylococcus aureus, auxotrophic for Ca-pantothenate, were isolated from the milk of four cows with mastitis. From one of these cows, a dwarf variant with an unknown metabolic disorder was isolated during the same period. Infections with auxo trophic staphylococci were always chronic and proved to be refractory to anti biotic treatment, though the causative micro-organism was highly susceptible to the drugs used as judged byin vitrotests.The authors acknowledge with appreciation the skilful help of Miss Nili Abramova.

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