Staphylococcus spp. - University of Mosul

Special Microbiology |Staphylococcus spp.| Dr. Ihsan Muneer Ahmed Page | 1

Key points • Gram-positive cocci in clusters resembling bunches of grapes • Grow on non-enriched media • Moderately-sized white or golden colonies • Facultative anaerobes, non-motile, catalase-positive • Commensals on mucous membranes and skin • Coagulase production correlates with pathogenicity • Comparatively stable in the environment • Cause pyogenic infections

Figure 1 Staphylococci in characteristic ‘bunches of grapes’ formations.

Ihsan Muneer Ahmed, BVMS, MSc, PhD

Lecturer, Department of Microbiology

College of Veterinary Medicine, University of Mosul, Mosul, Iraq

https://orcid.org/0000-0001-9698-702X

https://www.researchgate.net/profile/Ihsan_Ahmed3

Special Microbiology | 3rd

year 2020

1. Classification (According to Bergey’s Manual)

Phylum Firmicutes Class: Bacilli Order: Bacillales Family: Staphylococcaceae Genus: Staphylococcus

Staphylococcus aureus S. pseudintermedius S. intermedius S. aureus subsp. anaerobius S. chromogenes S. epidermidis S. equorum S. felis S. gallinarum S. saprophyticus

2. General characteristics

1. Staphylococci are Gram-positive cocci, approximately 1 μm in diameter, that tend to occur in irregular clusters resembling bunches of grapes (Fig. 1). The name derives from the Greek words staphyle and kokkos for a ‘bunch of grapes’ and a ‘berry’, respectively.

2. Staphylococcus species occur as commensals on skin and mucous membranes; some may act as opportunistic pathogens causing pyogenic infections.

3. Most staphylococci are facultative anaerobes and catalase-positive. They are non-motile and oxidase-negative and do not form spores.

4. A total of 43 species of Staphylococcus have been described to date some are coagulase positive (Table 1).

5. Coagulase production correlates with pathogenicity. On the other hand, some coagulase-negative staphylococci occasionally cause disease in animals and humans (Table 2).

Staphylococcus spp.

https://orcid.org/0000-0001-9698-702X

https://www.researchgate.net/profile/Ihsan_Ahmed3


Table 1 Coagulase-positive staphylococci and their clinical importance.

Species Hosts Clinical conditions

Staphylococcus aureusa Cattle Mastitis, udder impetigo

Sheep Mastitis

Tick pyaemia (lambs)

Benign folliculitis (lambs)

Dermatitis

Goats Mastitis

Dermatitis

Pigs Botryomycosis of mammary glands

Impetigo on mammary glands

Horses Scirrhous cord (botryomycosis of the spermatic cord), mastitis

Dogs, cats

Suppurative conditions similar to those caused by S. pseudintermedius

Poultry Arthritis and septicaemia in turkeys

Bumble foot

Omphalitis in chicks

S. pseudintermedius Dogs Pyoderma, endometritis, cystitis, otitis externa, and other suppurative conditions

Cats Various pyogenic conditions

Horses Rarely isolated

Cows Rarely isolated

S. intermedius Horses Isolated from nares

Pigeons

Isolated from upper respiratory tract

S. aureus subsp. anaerobius

Sheep Lymphadenitis

a, S. aureus can cause neonatal septicaemia and wound infections in many species. Table 2 Coagulase-negative staphylococci isolated from animals.

Species Host Source

S. chromogenes Cattle Milka

Pigs, poultry

Skin

S. epidermidis Cattle Milka

Dogs, horses

Wound infections

S. equorum Horses Skin

S. felis Cats Otitis externa, skin infections

S. gallinarum Poultry Skin infections

S. saprophyticus Cats Skin

Cattle Nostrils

a, occasionally isolated from cases of subclinical or clinical mastitis.


3. Usual habitat

Staphylococcal species occur worldwide as commensals on the skin of animals and humans. They are also found on mucous membranes of the upper respiratory tract and lower urogenital tract and as transients in the digestive tract.

Transfer of staphylococcal strains between animal species and between animals and humans is limited but of importance. Transfer of methicillin-resistant S. aureus strains from humans to animals or animals to humans is of particular significance.

4. Differentiation of Staphylococcus species

In clinical specimens, Staphylococcus species must be differentiated from Streptococcus species and from Micrococcus species (Table 3). Staphylococci are generally catalase-positive and streptococci catalase negative. Staphylococcus species are usually categorized by their colonial appearance, haemolytic pattern, biochemical profiles and ribosomal RNA gene restriction patterns.

Table 3 Differentiation of Gram-positive cocci.

a, oxidation-fermentation test, O oxidative, F fermentative.

1. Colonial characteristics: Staphylococcal colonies are usually white, opaque and up to mm in diameter. The colonies of bovine and human strains of S. aureus are golden yellow. Colonies of some coagulase-negative staphylococci are also pigmented.

2. Haemolysis in sheep or ox blood agar: Four staphylococcal haemolysins are recognized,

alpha, beta, gamma and delta. Individual haemolysins differ antigenically, biochemically and in their effects on the red blood cells of different animal species. Strains vary in their haemolysin-producing ability, and animal strains of S. aureus and S. pseudintermedius usually produce both alpha-haemolysin and beta-haemolysin. On ruminant blood agar, the alpha-haemolysin causes a narrow zone of complete haemolysis immediately around the colony, and the beta-haemolysin produces a wider zone of partial or incomplete haemolysis. This is referred to as double haemolysis (Fig. 2). These haemolysins act as toxins in vivo. Coagulase-negative staphylococci exhibit variation in their ability to produce haemolysis which usually develops slowly.

Organism Appearance in stained

smears

Coagulase production

Catalase production

Oxidase production

O-F testa

Bacitracin disc (0.04 units)

Staphylococcus spp. Irregular clusters

± + ‒ F Resistant

Micrococcus spp. Packets of four

‒ + + O Susceptible

Streptococcus and Enterococcus spp.

Chains ‒ ‒ ‒ F Resistant


Figure 2 The characteristic double haemolysis of S. aureus and S. pseudintermedius on

sheep or ox blood agar.

3. Slide and tube coagulase tests: In these tests, a suspension of staphylococci is mixed with rabbit plasma either on a slide or in a small tube. The fibrinogen in rabbit plasma is converted to fibrin by coagulase: – The slide test detects the presence of a bound coagulase or clumping factor on the

bacterial surface. A positive reaction is indicated by clumping of bacteria within 1 to 2 minutes.

– The tube test detects both free coagulase (staphylocoagulase), which is secreted by the bacteria into the plasma, and bound coagulase. It is the definitive test for coagulase production and a positive reaction is indicated by clot formation in the tube following incubation at 37°C for 24 hours.

4. Biochemical tests, which are commercially available, can be used to confirm the

staphylococcal species, although those currently available do not differentiate S. intermedius from S. pseudintermedius.

5. Molecular procedures such as the polymerase chain reaction are increasingly used by

diagnostic laboratories as well as research laboratories for definitive identification of staphylococcal species.

5. Pathogenesis and pathogenicity

Because staphylococci are pyogenic bacteria, they often cause suppurative lesions. Minor trauma or immunosuppression may predispose to the development of infection. As with many other bacterial pathogens, virulence attributes can be broadly classified into those that promote tissue colonization, immune evasion and tissue destruction.

1. Production of coagulase by staphylococci is an important indicator of pathogenicity. 2. Additional markers for pathogenicity are DNase activity and protein A production. 3. Biofilm formation by S. aureus, S epidermidis and S. pseudintermedius is a significant

virulence determinant and these species are capable of causing chronic prosthetic-device-related infections.

4. Enterotoxins produced by S. aureus are important causes of food poisoning in humans. S. pseudintermedius is also known to produce some similar toxins but the role of these toxins in food poisoning is not well defined. Virulence factors of S. aureus and their pathogenic effects are indicated in Table 4.


Table 4 Virulence factors, including toxins, of Staphylococcus aureus and their pathogenic effects.

Virulence factor Pathogenic effects

Coagulase Conversion of fibrinogen to fibrin. Fibrin deposition may shield staphylococci from phagocytic cells

Lipase, esterases, elastase, staphylokinase deoxyribonuclease, hyaluronidase, phospholipase

Enzymes which contribute to tissue destruction and virulence

Protein A Surface component which binds Fc portion of IgG and inhibits opsonization

Leukocidin Cytolytic destruction of phagocytes of some animal species

Alpha-toxin(alpha-haemolysin) The major toxin in gangrenous mastitis. It causes spasm of smooth muscle and is necrotizing and potentially lethal

Beta-toxin(beta-haemolysin) A sphingomyelinase which damages cell membranes

Exfoliative toxins Proteases which contribute to skin lesion development in humans, dogs and pigs

Enterotoxins Heat-stable toxins associated with staphylococcal food poisoning in humans

Toxic shock syndrome toxins(TSST)

Induce excessive lymphokine production, resulting in tissue damage. Bovine and human strains of S. aureus produce TSST-1. Sheep and goat strains produce a variant of this toxin. TSST-1 has superantigen activity

6. Diagnostic procedures

1. In suppurative conditions, the likelihood of staphylococcal infection must be considered and appropriate specimens such as exudates and mastitic milk collected for laboratory procedures.

2. Gram-stained smears of pus or other suitable specimens may reveal typical staphylococcal clusters.

3. Specimens are cultured on blood agar, selective blood agar and MacConkey agar and incubated aerobically at 37°C for 24 to 48 hours.

4. Identification criteria for isolates: – Colonial characteristics – Presence or absence of haemolysis – Absence of growth on MacConkey agar – Catalase production – Coagulase production – Biochemical profile – Molecular typing usually based on PCR procedures.


7. Methicillin-resistant staphylococcal infections in animals

1. Infection with Methicillin-resistant Staphylococcus aureus (MRSA) has been a major problem in human hospitals for many years but methicillin- resistant staphylococcal infections have become a major problem in veterinary medicine and animal production in the last decade.

2. Infection with MRSA in small animals, principally in dogs, has been recorded in many

countries, with wound infections, surgical site infections, pyoderma, otitis and urinary tract infections most commonly reported. Similar conditions have been reported in horses, including outbreaks of nosocomial infection in veterinary hospitals.

3. Transmission of infection between pets and humans, including veterinary personnel,

and between horses and humans has been reported.

Staphylococcus spp. - University of Mosul

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