Introduction field strains of this pathogen are considered hot points of food inspection related with specific control of milk.
1. Introduction field strains of this pathogen are considered hot points of food inspection related with specific control of milk.
There are many cow farms producing milk in Fieri Raw milk produced by cows at the temperature of region, Albania. These farms are producing almost cow’s body is considered as one of the best mediums 30% of the total cow milk production. Fresh milk is for growth of Staphylococcus aureus [1, 2]. Milk and mostly sold in Fieri and other cities including the its products are common vehicles of Staphylococcus capital city, Tirana. The other quantity of the produced aureus transmission to humans. S. aureus as milk is processed in three big processing milk foodborne pathogen causes different infections in factories located in this region. Considering raw milk humans and animals presenting themselves with of cows produced as main food used by many severe syndromes in skin and other diseases such as consumers in Albania, control for presence of pneumonia and septicemia [3, 4]. In dairy cattle, S. coagulase positive S. aureus and antibiotic resistance of aureus is frequently associated with subclinical
Corresponding author: Kapllan Sulaj, Ph.D., professor, mastitis and may contaminate milk and other dairy
research fields: veterinary microbiology, food safety. E-mail: products [2, 5, 6]. The mammary glands serve as a [email protected].
Incidence of Coagulase Positive Staphylococcus aureus in Raw
Cow Milk Produced by Cattle Farms in Fieri Region in Albania
source of infection, when they are infected by S. mL) were collected aseptically from each cow in aureus contaminating at the same time cows and the
different farms after the milking process. At the end of milk. Contaminated milking equipment and the hands
the milking procedure, triplicate bulk tank milk of the milkers are also common vehicles for samples (100 mL) were also collected in sterile glass transmission of St. aureus infection [7, 8].
flasks. One hundred five samples of milk from Different strains of S. aureus are producing several
individual cow’s milk were aseptically collected, and staphylococcal virulence factors, including transported to the laboratory in coolers with ice enterotoxins (SEA to SEE and SEG to SEQ). Other
(4-8 ºC), and analyzed on the same day. toxins, such as exfoliative toxin A and B, and toxic
2.2 Isolation, Identification and Enumeration of S. shock syndrome toxin (TSST-1) are identified as
aureus
responsible causes in intoxication [9]. Staphylococcal food is recognized as a cause of food-borne diseases.
Bacteriological methods used for isolation and Even though pasteurization kills S. aureus cells,
identification of S. aureus were performed according thermostable SEs generally retain their biological
to ISO 6888-1, 1994. Serial dilutions of samples were activity. This risk caused by contaminated raw milk of
prepared according to IS0 6887-1, 1994. Samples cows with S. aureus can be increased by adequate
were spread on Baird Parker agar (Oxoid) pasteurized milk products and by adequate chilling of
supplemented with egg yolk and tellurite emulsion
raw milk until heat treatment, followed by very strict (1%) (Oxoid). From each serial dilution (10 , 10 and preventive measures [8]. Incidence of St. aureus in -5 10 ) were transferred, with sterile pipette 0, 1 mL of
-3 -4
raw milk samples is reported in many cases and at a the test sample (milk) to each of two Baird Parker agar variety of values [4, 8]. Food intoxications caused by -2 plates. This process is repeated for the 10 dilution
using raw milk are well known and the majority holds and for further decimal dilutions. Carefully inoculums S. aureus or its enterotoxins as responsible agents
of samples were spread as quickly as possible over the [7, 8].
surface of the Baird Parker agar plates trying not to Prevalence of resistance for antimicrobial agents
touch the sides of the dish, using the spreader. Agar among bulk milk and composite milk Staphylococcus o plates prepared were incubated at 37
C for 24-48 h. aureus is reported with average values ranging from
All inoculated plates were controlled for presence of 12% to 25%, compared to 49.3% and 69.5% among
coagulase positive staphylococci differentiating isolates from human anterior nares and hand swabs,
typical and atypical colonies according to specific respectively [2, 6, 10]. Resistance to penicillin,
characteristics. Colonies suggestive colonies of S. vancomycin and methicilin is the highest among other
aureus were submitted for the following tests antibiotics [9, 11]. Bulk milk containing relatively
performed using API Staph strips (bio Merieux) as high counts of enterotoxigenic S. aureus may well as hemolysis, coagulase and catalase tests. constitute health hazard to consumers.
Microscopic and biochemical test were completed according to the method applied. Enumeration of St.
2. Materials and Method
aureus was achieved belonging to above ISO method
2.1 Sampling applied. In this case a value found was raised by a
factor of 10 because milk samples are liquid. The study was conducted in dairy farms located in the region of Fieri in Albania from 2007 to 2009. On 2.3 Antibiotic Resistance (Penicillin, Methicilin,
each farm, individual milk samples were obtained Vancomycin) from all cows selected for this control. Samples (300
Isolated colonies of S. aureus with the same
Incidence of Coagulase Positive Staphylococcus aureus in Raw
Cow Milk Produced by Cattle Farms in Fieri Region in Albania
morphological characteristics were selected from a in Fieri region is 18% (19/105). From 2007 to 2008 primary Baird Parker agar plate culture and the incidence was confirmed in 14% of analyzed transferred into a tube containing 4-5 mL of broth
samples. The higher value of presence of this medium. Tubes were incubated at 35 °C for 2-6 h and
pathogen has been evaluated from 2008 to 2009 in then applied with a sterile cotton swab on a dried
22% of raw milk samples controlled in this year. An surface of a Mueller-Hinton and blood agar plates.
analogue study confirmed the value 14.7% of the Anti-microbial disks were dispensed onto the surface
incidence of Staphylococcus aureus in raw milk of the inoculated agar plate. Each disk was pressed
samples [11]. Somehow, this incidence is higher or down to ensure complete contact with the agar surface.
lower depending on the health status of the milking Disks (Oxoid) of penicilln, methicillin and cows [1-10, 12-16]. Postpartum mastistis and vancomycin were distributed evenly no closer than 24
subclinical cases are indicated in found values mm from center to center. The plates were inverted
reported by many studies [16]. From our study, it is and placed in an incubator at 35 °C. After 16-18 h of
clearly showed that coagulase positive Staphylococcus incubation, each plate was examined. The resulting
aureus isolates are more atypical strains than typical. zone of inhibition (halo) with uniformly circular and
This fact is mentioned also by other authors who confluent lawn of growth was considered as resistant.
described that in raw milk samples, the number of The diameters of the zone of complete inhibition were
coagulase positive S. aureus atypical isolates is higher measured using sliding caliber, including the diameter
than strains appearing typical characteristics of growth of the disk. Interpretation of the results was performed
colonies in Baird Parker and blood agar plates [15]. according to specific tables in which the values range
Atypical strains of Staphylococcus aureus are reported for: resistance, intermediate and sensibility to used
as more virulent and they can produce entertoxines antibiotics.
types A and B which cause very strong intoxication in humans and calves [6]. However, a comparison of