aureus Staphylococci on Staphylococcus aureus Growth and ...

In Vitro Effect of Bovine-associated Non-

aureus Staphylococci on Staphylococcus

aureus Growth and RnaIII Expression

M-teamUGent and Mastitis and Milk Quality Research Unit

National Mastitis Council

60th Annual Meeting

Drs. Bruno Toledo Silva

Dr. Fernando Nogueira de Souza

Prof. dr. Sofie Piepers

Prof. dr. Sarne De Vliegher

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DISTRIBUTIONNon-aureus staphylococci

INTRODUCTION


DISTRIBUTION

Mastitis??

INTRODUCTION

Non-aureus staphylococci


NAS??

INTRODUCTION

ROLE

• In vitro growth inhibition of major mastitis pathogens (De Vliegher et al., 2004; Braem et al., 2014);

• In vivo protection against IMI when colonizing TA and mammary gland (Piepers et al., 2010, 2011);

• NAS influences the expression of S. aureus virulence factors controlled by the quorum sensing system (Mahmmod

et al., 2018).

• IMI (Piessens et al., 2011; Supré et al., 2011; Fry et al., 2014);

• ↑ SCC (Tomazi et al., 2015; Valckenier et al., 2019).;

• ↑ SCC , but no impact milk yield (Valckenier et al., 2020);


BACTERIAL COMMUNICATION

Bacterial infections

Cell-cell communication

Quorum Sensing

INTRODUCTION


QUORUM SENSING (QS)

QS

INTRODUCTION


STAPHYLOCOCCUS AUREUS

o Accessory gene regulator (agr) system

Genes

o RNAIII

o ↑ hla

o ↓ spa

NAS x S. aureusinteractions

o Niche competitiono Colonization

?

(Novick et al., 1993; Quave & Horswill, 2014)

INTRODUCTION


Milk

Teat Apex

Influence

S. aureus agr systemNAS

OBJECTIVE


NAS ISOLATES

NAS species Milk1 TA Total

Staphylococcus chromogenes 28 6 34

Staphylococcus epidermidis 7 4 11

Staphylococcus simulans 10 4 14

Total 45 14 59

1Piessens et al. (2011)

MATERIAL & METHODS


BETA GALACTOSIDASE LIQUID ASSAY

S. aureusrnaIII::LacZ

Overnight cultures

1:1

CFU/mL

OD activity

59 NAS S. chromogenes

S. simulans

S. epidermidis

4h

MATERIAL & METHODS


Milk

Teat Apex

Influence

S. aureus agr systemNAS

MATERIAL & METHODS


BETA GALACTOSIDASE LIQUID

RESULTS

Final linear mixed effect model for the β-galactosidase liquid assay for rnaIII gene expression including the positive and negative controls


BETA GALACTOSIDASE LIQUID

RESULTS

Final linear mixed regression model for the β-galactosidase liquid assay for rnaIII gene expression and final model corrected for in vitro growth inhibition of Staphylococcus aureus by bovine non-aureus staphylococci (NAS)

With in vitro growth inhibition Without in vitro growth inhibition


❑ NAS species isolated from milk and teat apices of dairy cowsinterfered with S. aureus agr activation– Variable effect

– Species-dependent

– Habitat-independent

– Not dependent on in vitro growth inhibition

❑ Antimicrobial compounds produced by NAS, which have yet to beidentified (protein, non-protein and NAS cells themselves - data notyet published), might substantially reduce rnaIII gene expression inS. aureus in ways that are consistent with repression of the agrsystem

❑ The biological importance of these interactions in the bovinemammary gland remains still unclear, hence further investigationsare needed

DISCUSSION

RNAIII EXPRESSION

(Mahmmod et al.; 2018)


CONCLUSIONS

❑ Most of the NAS isolates were able to influencethe agr quorum sensing system of S. aureus at some level

❑ This effect seems to be species-dependent, but not habitatdependent

❑ There was no association between the capacity of NAS toinhibit the growth of S. aureus in vitro and to repress thequorum sensing system, although thischaracteristic weakened the regulation effect of NAS specieson S. aureus agr

❑ Other mechanism, rather than in vitro growth inhibition,possibly plays a role in the regulation of S. aureus agr

aureus Staphylococci on Staphylococcus aureus Growth and ...

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