Review of infections by Staphylococcus aureus in the dairy cow

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Review of infections by Staphylococcus aureus in
the dairy cow

• Dr. Gaddo Vicenzoni
• Via S. Giacomo, 5 - 37135 Verona
• Tel. [phone] - Fax [phone]
• E-mail gaddovic.izsv_at_interbusiness.it
• Reggio Emilia, Italy, 17.12.99

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LEGAL REGULATIONS characteristics of raw milk
as of 01.01.98 Presidential Decree 54/97 (Italy)

• Content of microorganisms (per ml) lt 100,000
• Cell content (per ml) lt 400,000
• Staphylococcus aureus (per ml)
• m 500 M 2,000
• no. 5 c
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Definition of the Problem

• QUALITY OF THE MILK
• SOMATIC CELLS
• MASTITIS
• INFECTIONS

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Epidemiological classification of the
microorganisms

• Contagious
• S. agalactiae
• Staph. aureus
• C. bovis
• Mycoplasmas

• Environmental
• Non-agalactiae streptococci
• Coagulase-negative staphylococci
• Enterobacteria

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Epidemiology of mastitis due to Staphylococcus
aureus
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Control of infections by Staphylococcus aureus

• TOTAL COMMITMENT OF THE BREEDER
• HEALTH AND HYGIENE MEASURES
• IDENTIFICATION INFECTED ANIMALS
• THERAPY

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THE BREEDER MUST BE INFORMED OF

• REGULATORY ASPECTS
• ECONOMIC ADVANTAGES
• COMMITMENT REQUIRED

ALL CLEARLY AND SIMPLY THE VETERINARIAN
MUST BE A LEADER
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Control of infections by Staphylococcus aureus

• TOTAL COMMITMENT OF THE BREEDER
• HEALTH AND HYGIENE MEASURES
• IDENTIFICATION OF INFECTED ANIMALS
• THERAPY

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HEALTH AND HYGIENE MEASURES

• CONTROL OF SOURCES OF INFECTION
• MILKING PRELIMINARIES
• RATIONAL MILKING
• NUTRITION OF HEIFERS
• QUARANTINE OF NEW ARRIVALS

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CONTROL OF SOURCES OF INFECTION - ENVIRONMENT
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CONTROL OF SOURCES OF INFECTION ANIMAL
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HEALTH AND HYGIENE MEASURES

• CONTROL OF SOURCES OF INFECTION
• MILKING PRELIMINARIES
• RATIONAL MILKING
• NUTRITION OF HEIFERS
• QUARANTINE OF NEW ARRIVALS

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PRE-MILKING DISINFECTION efficacy against
contagious mycoplasmas
new infections
Source USA
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HEALTH AND HYGIENE MEASURES

• CONTROL OF SOURCES OF INFECTION
• MILKING PRELIMINARIES
• RATIONAL MILKING
• NUTRITION OF HEIFERS
• QUARANTINE OF NEW ARRIVALS

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RATIONAL MILKING
control of milk cleanliness and
milking attachment to milking group
milking machine dripping dipping
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Prevention of new infections by use of
post-dipping (experimental trials)

• Staphylococcus aureus
• iodophore 1.0
• chlorhexidine 0.5

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HEALTH AND HYGIENE MEASURES

• CONTROL OF SOURCES OF INFECTION
• MILKING PRELIMINARIES
• RATIONAL MILKING
• NUTRITION OF HEIFERS
• QUARANTINE OF NEW ARRIVALS

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HACCP - contagious microorganisms
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Control of infections by Staphylococcus aureus

• TOTAL COMMITMENT OF THE BREEDER
• HEALTH AND HYGIENE MEASURES
• IDENTIFICATION OF INFECTED
• ANIMALS
• THERAPY

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IDENTIFICATION OF INFECTED ANIMALS

• BACTERIOLOGICAL EXAMINATION
• CYTOLOGICAL EXAMINATION

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PROBABILITY OF HAVING A SAMPLE POSITIVE FOR S.
AUREUS FROM AN INFECTED QUARTER ON THE BASIS OF
THE NUMBER OF SAMPLES
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STAPH. AUREUS WITH LOW SHEDDING CYCLE
CFU/ml
no. of milkings after inoculation
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Probability of having growth of 1 or more
colonies of Staph. aureus
Buelow Nordlund NMC (1999),68-75
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IDENTIFICATION OF INFECTED ANIMALS

• BACTERIOLOGICAL EXAMINATION
• CYTOLOGICAL EXAMINATION

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SCC CHOICE OF THRESHOLD VALUE

HEALTHY 50 INF. 50
SENS. 0.90 SPEC. 0.80 PV 0.82 PV -
0.89
TN
cows
TP
FN
FP
linear score
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SCC CHOICE OF THRESHOLD VALUE

HEALTHY 50 INF. 50 SENS. 0.80 SPEC.
0.90 VP 0.89 VP - 0.82
cows
TN
TP
FN
FP
linear score
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SCC CHOICE OF THRESHOLD VALUE

HEALTHY 50 INF. 50 SENS. 0.65 SPEC.
0.95 VP 0.93 VP - 0.73
cows
TN
TP
FN
FP
linear score
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Control of infections by Staphylococcus aureus

• TOTAL COMMITMENT OF THE BREEDER
• HEALTH AND HYGIENE MEASURES
• IDENTIFICATION OF INFECTED ANIMALS
• THERAPY

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IN VITRO SENSITIVITY AGAINST STAPH. AUREUS

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MIC of strains of Staph. aureus
Watts JL- Salomom S.A. J.Dairy Sc. (1997)
80788-791(adapted)
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Calculation of the species-related MIC for
Rifaximin
MIC (?g/ml)
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Lactation therapy against Staphylococcus aureus

Owens et al J.Dairy Sc. (1997) 80313-317
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RESULTS OF DRY-COW THERAPY AGAINST S. AUREUS
WITH RIFAXIMIN
76.7
23.3
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In spite of everything, Staph. aureus remains the
principal pathogen of mastitis in dairy cows
throughout the world. All agree on the need to
improve our epidemiological and pathogenic
knowledge of this microorganism
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Objective

• To identify molecular typification methods for
  strains of Staphylococcus aureus isolated from
  cases of bovine mastitis
• To evaluate, using objective methods, any
  correlations between clusters and virulence of
  strains

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Molecular typification

• X region of the Spa gene

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Mechanisms of inhibition of phagocytosis by
protein A
FcR
F(c)
PA
PA
F(ab)
S. aureus
PMN
PA
F(c)
F(ab)
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Gene of Protein A (Spa)
Rsa I GT/AC


Rsa I
X region
Spa1 5 - TGTAAAACGACGGCCAGTGCTAAAAAGCTAAACGATG
C - 3 Spa2 5 CAGGAAACAGCTATGACCCCACCAAATACA
GTTGTACC 3
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ANALYSIS OF THE X REGION USING PCR
2 4 5 6 7 8 9 10
11 12 M
213 bp
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DISTRIBUTION OF STRAINS OF STAPH. AUREUS ON THE
BASIS OF THE NUMBER OF REPETITIONS

No. of repetitions
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Virulence of the strain

• Measurement of phagocytic activity of PMN using
  chemoluminescence

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Activation of the oxidative burst in neutrophils
Opsonization (complement, Ab)
1-anchorage
glucose
O2
NADP
3-endocytosis
HMP
NADPH Oxidase
O2
NADPH
2-activation
G-6-P
excitation
H2O
2
OH-
O2
MPO
OCl-
Background state
4-killing
light
5-exocytosis
LUMINOL
chemoluminescence
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Determination of phagocytic activity

• preparation of bacterial suspension
• opsonization
• isolation of blood leucocytes
• measurement of luminol-amplified phagocytic
  activity

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Distribution of phagocytic activity per number of
repetitions
P 0.0501
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Distribution of phagocytic activity per number of
repetitions, using the generalised linear model
RLU
no. of repetitions
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Distribution of phagocytic activity per number of
repetitions, using the generalised linear model
RLU
P 0.0226
? 7
? 8
no. of repetitions
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Conclusion

• Strains of Staph. aureus with a high number of
  repetitions regarding the X region of the gene
  Spa are able to induce, in vitro, a less intense
  phagocytic response and can thus elude the
  specific defences of the udder.