Classification and biological characteristics of coagulase-positive Staphylococci isolated from animals

A total of 165 coagulase-positive staphylococcal strains of different origin (142 S. aureus strains and 23 S. intermedius strains) were subjected to biological typing in accordance with the schemes of Hajek-Marsalek and Meyer-Witte. The former of these schemes permitted to identify 68% and the latter 18% of S. aureus strains. The cultures isolated from swine and chickens had the most uniform composition: 85-86% of the strains belonged to biotype B. 44% of the strains isolated from cows and sheep belonged to biotypes C (ecovars bovis and ovis) and A (ecovar hominis); the rest of the strains could not be identified. 96% of the strains isolated from minks were made up of S. intermedius, more than a half of them belonging to biotype E (ecovar canis). In 80% of S. aureus strains and 48% S. intermedius cultures protein A was detected. Only 9% of S. aureus strains of animal origin were found capable of producing enterotoxins (A-D). The expediency of working out a unified scheme for the biotyping of coagulase-positive staphylococci is discussed.     

Resistance to antibiotics and heavy metal salts of coagulase-positive staphylococci belonging to different biological types (ecovars)

A total of 189 strains of S. aureus isolated from cows, sheep, swines, poultry, monkeys, rabbits, foxes, and humans and 23 strains of S. intermedius isolated from minks and sables were studied. The staphylococci belonged to different biological types (according to the Hajek-Marsalek's scheme) and ecovars (according to the Meyer-Witte's scheme). The strains were studied with respect to their resistance to 10 antibiotics (benzylpenicillin, streptomycin, tetracycline, levomycetin, erythromycin, oleandomycin, neomycin, kanamycin, monomycin, and novobiocin), mercuric chloride and cadmium sulfate. As a whole the frequency of resistance to the above preparations among the staphylococci of the animal origin was not high. Differences in the frequency and range of the resistance between strains belonging to different biological types (ecovars) were shown. The highest number of the resistant cultures was detected among the strains of the biological type E (ecovar canis) and atypical strains of S. intermedius isolated from the minks. The least number of the resistant cultures was detected among the strains of the biological type C1 (ecovar bovis) isolated from the cows. It was found that almost all strains of S. intermedius were resistant to cadmium sulfate. This may be used as an additional characteristic of the species.     

Frequency of the isolation of staphylococci from domestic animals and strain identification

Staphylococci occur in donkeys more frequently than in other animals, and only from donkeys coagulase-negative staphylococci, characteristic of humans (S. hominis, S. capitis, S. cohnii), were isolated. Least frequently staphylococcal carrier state was registered in cats; in these animals only coagulase-negative strains were found to occur. From 30 donkeys coagulase-positive staphylococci belonging to 47 S. aureus strains were isolated. These strains differed from known ecological variants in their biological properties, thus suggesting the existence of S. aureus ecovar specific for donkeys. These strains did not coagulate human, bovine and ovine plasma, but coagulated rabbit plasma in 100% of cases and donkey plasma only in 53% of cases; at the same time they relatively often produced delta hemolysin, rarely phosphatase and hyaluronidase and never fibrinolysin. These strains were typed by KPC phages, mainly 116 and 117.     

Phage typing of the coagulase-positive staphylococci isolated from various species of animals

More than 200 coagulase-positive strains of animal origin have been studied by means of Staphylococcus aureus typing phages, belonging to two international sets and intended for typing staphylococci isolated from large cattle and humans, and experimental "chicken" phage A 1591. Among S. aureus strains the cultures isolated from swine, cows, chickens, and belonging to biotypes B1, C1, B2, respectively, have been mostly (in 78.5-90.0% of cases) determined by phage typing. The strains belonging to one biotype have proved to be sensitive predominantly to the same phages. In this connection further differentiation of staphylococci within individual biotypes by means of the phages used in these experiments seems to be impracticable. S. intermedius strains have been found to be completely resistant to the above phages, which confirms that S. intermedius is rightly considered to be an independent species of coagulase-positive staphylococci.     

Antiseptic resistance of clinical strains of Staphylococcus aureus

To evaluate the activity of antiseptics and the sensitivity-resistance of bacteria to antiseptics, a number of characteristics has been used, including the minimum inhibiting concentration for different strains, the frequency of statistical and clinical resistance, the antiseptic activity index. A wide spread of S. aureus strains isolated from patients with hospital infections has been revealed. Differences in the resistance of bacterial strains have been established, depending on the type of the antiseptic and the ecovar of bacteria: among hospital ecovars, resistant strains occur more frequently and can resist a wider range of antibiotics. In staphylococcal hospital ecovars the occurrence and level of resistance to a number of widely used antiseptics increase with time. In connection with a wide spread of staphylococcal hospital strains resistant to antiseptics, measures on the control of the circulation of such strains should be introduced into hospitals, and the data thus obtained should be used for the periodic reevaluation of antiseptics used in medical practice and for the choice of preparations to be used for individual therapeutic and prophylactic antisepsis.     

Thermal Resistance of Staphylococcus aureus in Milk, Whey, and Phosphate Buffer

The thermal resistance of four strains of coagulase-positive Staphylococcus aureus was determined in phosphate buffer, whole milk, skim milk, and Cheddar cheese whey. The logarithmic order of death prevailed until about 99.99 to 99.999% of the organisms were destroyed, after which there was a decline in the rate of destruction. The organisms were more resistant in skim milk and Cheddar cheese whey than in phosphate buffer and whole milk. Thermal resistance varied among strains of S. aureus but was consistent with individual strains. As the age of cultures of strain B-120 increased from 12 to 228 hr, the D(55) values increased from 0.95 to 3.0. The thermal resistance of cultures obtained from survivors to partial thermal destruction was similar to that of the parent cultures.     

A simplified system for biotyping Staphylococcus aureus strains isolated from different animal species

D evriese , L.A. 1984. A simplified system for biotyping Staphylococcus aureus strains isolated from different animal species. Journal of Applied Bacteriology 56 , 215–220.
A biotyping system for Staphylococcus aureus strains is proposed which is a simplified version of biotyping procedures described in the literature. It differentiates Staph. aureus strains from man and animals into host-specific ecovars and biotypes which are not host-specific. With the help of tests for βhaemolysin, staphylokinase, coagulation of bovine plasma and the crystal-violet reaction, the origin of many but not all Staph. aureus strains can be determined: 604 of 809 strains from man. poultry, cattle, pigs, goats, rabbits and foods could be alloted to four ecovars which are typically associated with man, poultry, sheep and goats and cattle. The other strains belonged to five non-host specific biotypes.     

Antagonistic activities of coagulase-positive staphylococci

Antagonistic activities were investigated by Frédérioq's plate method in 1,014 coagulase-positive staphylococcal strains of different species and origins. Staphylococcin effect was demonstrated in 12 (2%) of the 559 Staphylococcus aureus strains, in 51 (18%) of the 283 S. intermedius strains, and in 1 (3%) of the 36 S. hyicus subsp. chromogenes strains. Lysostaphin was identified in the 15 (5%) S. intermedius strains. In addition, heat-stable bacteriostatic agent was detected in 45 (33%) and heat-labile bacteriolytic agent in 7 (5%) of the 136 S. hyicus subsp. hyicus strains. An attempt was made at antagonistic activity typing in all of the active staphylococcal strains.     

Staphylococci in the microbiocenosis of the skin of the hands]

The skin autoflora on the pad of a forefinger and the back and palm of a hand was studied in 40 healthy males aged 18-60 years by the modified washing and scraping method of P. Williamson and A. Kligman. 638 cultures of aerobic microorganisms, including coccal (55.3%) and bacilliform (44.7%) microbes, were isolated. In 6 persons (15%) coagulase-positive staphylococci were detected. Out of 10 coagulase negative species of this genus, S. epidermidis, S. saprophyticus and S. warneri occurred most frequently on the skin of hands. The highest density of bacterial populations (10.970 +/- +/- 1.845 cells/sq. cm) was registered on the back of hands, the surface of palms was found to have somewhat lower density (8.679 +/- 1.282 cells/sq. sm) and the skin of forefingers, the lowest density of bacterial populations (6.878 +/- +/- 1.137 cells/sq. sm). 17.5% of examined persons were found to be carriers with S. aureus isolated from their nasal mucosa. S. aureus isolated from the skin surface and the nasal cavity of different persons belonged to different phage variants, but S. aureus isolated from the nasal cavity and the skin of the same person belonged to one phage variant.     

Characteristics of staphylococci isolated from man, poultry and some other animals

Of 281 strains of staphylococci isolated from man and animals 36 (12.8%) were coagulase-positive and 245 (87.2%) were coagulase-negative. Staphylococcus aureus and Staph. intermedius were the commonest coagulase-positive staphylococci isolated from the hosts examined. Of the 20 strains that remained unclassifiable, 14 were isolated from sheep and goats.     

Characterization of coagulase-positive staphylococci isolated from free-living birds.

A total of 129 coagulase-positive staphylococcal strains were isolated from the nasopharyngeal region of free-living birds--81 (16%) from 501 birds of prey, 29 (25%) from 117 water birds, and 19 (2%) from 937 pheasants. Staphylococcus aureus and Staphylococcus intermedius were identified in 64 (79%) and 17 (21%) strains from birds of prey, in 1 (3%) and 28 (97%) strains from water birds, and in 4 (21%) and 15 (79%) strains from pheasants, respectively. Of the total number of the 69 S.aureus strains 45 (65%) could be biotyped. Biotype D prevailed in the strains from birds of prey. Among all the 60 S.intermedius strains 41 (68%) biotype 1 and 19 (32%) biotype 3 strains could be recognized. Biotype 1 predominated in strains from water birds and pheasants, while biotype 3 in strains from birds of prey. Resistance to antibiotics was recorded in 8 (12%) S.aureus and 5 (8%) S.intermedius strains only. Fifty-six (81%) of the 69 S.aureus strains could be typed with human phages, 30 (44%) with bovine and 41 (51%) with chicken phages. It is evident that the host range of Shimizu's (CH) phages involves not only S.aureus from chicken but also from any other birds. None of these strains was typable with canine phages. Out of the 60 S.intermedius strains 45 (75%) were lysed with Blouse and Meekins' canine phages. When human, bovine, and chicken phages were used, all the 60 strains were completely resistant.     

PCR detection of staphylococcal enterotoxin genes in Staphylococcus spp. strains isolated from meat and dairy products. Evidence for new variants of seG and seI in S. aureus AB-8802

AIMS: Evaluation of the occurrence of most known staphylococcal enterotoxin (SE) genes, egc (enterotoxin gene cluster) and TSST1 (toxic shock syndrome toxin 1) gene in both coagulase-positive (CPS) and coagulase-negative (CNS) staphylococcal strains isolated from meat and dairy products. METHODS AND RESULTS: Specificity and reliability of the PCR detection methods used were ascertained by using nine reference strains of Staphylococcus (S. aureus) harbouring SE genes (seA to seE; seG, seH, seI, seM, seJ, seN and seO) and egc (containing the following sequence of genes: seO, seM, seI, phient1, phient2, seN and seG). Of 109 wild Staphylococcus spp. strains analysed, only 11 S. aureus strains were SE and/or TSST1 PCR-positive. The last 11 strains also appeared to harbour the egc. Restriction endonuclease analysis of part of the egc of both reference and wild strains showed that different variants of the egc exist. Moreover, nucleotide sequences of seG and seI indicate that the egc of the strain AB-8802 is characterized by the presence of variants of these enterotoxins (seGv and seIv). CONCLUSIONS: The occurrence of SE genes in CNS and other non-S. aureus species isolated from Napoli-type salami, raw water buffalo milk and natural whey cultures used for mozzarella cheese manufacturing is very rare. SIGNIFICANCE AND IMPACT OF THE STUDY: During this study it was shown that at least five different egc may exist in S. aureus. A thorough study of egc polymorphism should provide further insight into the phylogenetics of the egc.     

Characteristics of staphylococci isolated from man, poultry and some other animals

Of 281 strains of staphylococci isolated from man and animals 36 (12–8%) were coagulase-positive and 245 (87–2%) were coagulase-negative. Staphylococcus aureus and Staph. intermedius were the commonest coagulase-positive staphylococci isolated from the hosts examined. Of the 20 strains that remained unclassifiable, 14 were isolated from sheep and goats.     

Comparison of Vogel-Johnson and Baird-Parker media for membrane filtration recovery of staphylococci in swimming pool water

Previous studies have indicated that the coagulase-positive Staphylococcus (Staphylococcus aureus) has potential as a useful indicator of the infection hazard associated with the use of swimming pools and other recreational waters. However, before this indicator system can be used effectively, a recovery system that is sufficiently selective, accurate, and reliable for the enumeration of S. aureus must be developed. In this study, Vogel-Johnson (VJ) and Baird-Parker (BP) agars were compared for efficacy in the primary isolation and recovery of S. aureus from swimming pool water. For equal sample volumes of pool water containing adequate free chlorine residual, VJ agar was found to be more selective for staphylococcal species and less inhibitory to general cell growth than was BP agar. However, neither medium was found to be sufficiently differential to permit the accurate identification of S. aureus. In contrast, water samples obtained from a swimming pool containing very low levels of chlorine (none of which was in the free form) showed abundant growth of staphylococci on both test media, with both VJ and BP agars showing increased sensitivity for the detection of S. aureus. Thus, VJ and BP agars show increased sensitivity for the detection of coagulase-positive staphylococci from unchlorinated versus chlorinated waters.     

Comparison of Vogel-Johnson and Baird-Parker media for membrane filtration recovery of staphylococci in swimming pool water.

Previous studies have indicated that the coagulase-positive Staphylococcus (Staphylococcus aureus) has potential as a useful indicator of the infection hazard associated with the use of swimming pools and other recreational waters. However, before this indicator system can be used effectively, a recovery system that is sufficiently selective, accurate, and reliable for the enumeration of S. aureus must be developed. In this study, Vogel-Johnson (VJ) and Baird-Parker (BP) agars were compared for efficacy in the primary isolation and recovery of S. aureus from swimming pool water. For equal sample volumes of pool water containing adequate free chlorine residual, VJ agar was found to be more selective for staphylococcal species and less inhibitory to general cell growth than was BP agar. However, neither medium was found to be sufficiently differential to permit the accurate identification of S. aureus. In contrast, water samples obtained from a swimming pool containing very low levels of chlorine (none of which was in the free form) showed abundant growth of staphylococci on both test media, with both VJ and BP agars showing increased sensitivity for the detection of S. aureus. Thus, VJ and BP agars show increased sensitivity for the detection of coagulase-positive staphylococci from unchlorinated versus chlorinated waters.     

Characterization of yellow-pigmented and motile enterococci isolated from intestines of the garden snail Helix aspersa

AIMS: Enterococci associated with garden snails (Helix aspersa) were studied in order to obtain reliable species identification and characterization. METHODS AND RESULTS: Twelve yellow-pigmented and motile enterococci, isolated from the intestines of garden snails, were phenotypically close to Enterococcus casseliflavus, but they showed certain unusual biochemical characteristics. tRNA intergenic length polymorphism analysis (tDNA-PCR) divided all strains studied into two groups, in full agreement with biochemical test results. 16S rDNA sequencing, DNA base composition analysis and DNA-DNA hybridization results showed unambiguously that the enterococci studied belonged to the species Ent. casseliflavus. The representative strains of described ecovars were deposited in the Czech Collection of Microorganisms (CCM) as Ent. casseliflavus CCM 4868, 4869, 4870 and 4871. CONCLUSIONS: Enterococcus casseliflavus associated with garden snails can be subdivided into groups. SIGNIFICANCE AND IMPACT OF THE STUDY: Enterococcus casseliflavus differs from other enterococcal species in that it is typically associated with plants, soil, water and invertebrate animals. The different groups that can be found in these widely occurring bacteria are possibly source-specific ecovars, as exemplified by the Ent. casseliflavus inhabiting the intestines of snails.     

RAPD-PCR analysis of Staphylococcus aureus strains isolated from bovine and human hosts

Staphylococcus aureus is one of the most important pathogens in humans and animals. In this study eighty strains were analyzed by RAPD-PCR to assess the genetic relationship between S. aureus isolates from bovine and human hosts. Results were compared with those obtained by biotyping. Fifty-two percent of the S. aureus isolates belonged to a host specific biotype (human, bovine and poultry). Bovine and human ecovars were the most prevalent. Dendrogram obtained by RAPD results showed that all the isolates clustered into eleven groups (A-K) at a relative genetic similarity of less than 30% when analyzed with the three primers. Group A clustered 95% of the human host isolates and the remaining groups (B-K) clustered the bovine host isolates. Principal coordinate analysis also showed that the isolates could be arbitrarily divided into two groups, bovine and human, by the second coordinate. Only 9 isolates (11%) were not clustered into these groups. The genetic diversity among the S. aureus isolates from bovine hosts is relatively low compared to that of isolates from human hosts. There were no statistically significant differences among isolated from bovine and human hosts. This study shows that RAPD-PCR assayed with three primers can be successfully applied to assess the genetic relationship of S. aureus isolates from different hosts.     

Bacteriocinogenic properties of Staphylococci isolated from the oral cavity

Two hundred ninety two staphylococcal strains were isolated out of 130 saliva samples taken from children and adults, among which 116 were coagulase-positive and 176 coagulase negative. Bacteriocinogenic activity against Staphylococcus aureus strain Oxford 209P was found in 13 (4.5%) of the strains only. On the other hand, when a set of 15 sensitive staphylococcal strains selected by cross checking was used for the study 260 (89.0%) strains were found to be bacteriocinogenic. It was found that a higher percentage of coagulase-positive staphylococcal strains is sensitive to staphylococcins than of coagulase-negative strains. However, mean zone of inhibition is smaller in the case of former than of the latter strains. It was shown, that in the case of active strains a positive correlation exists between a percentage of coagulase-positive and negative strains inhibited by them and also between a percentage of all inhibited strains and a mean diameter if the growth inhibition zone. Simultaneous occurrence in saliva of two or more staphylococcal strains was found in 106 persons examined. In 93.4% of those cases coexisting strains did not show antagonistic properties: in remaining 6.6% despite of the number of simultaneously existing strains in oral cavity only one strain showed antagonistic properties against the remaining strains.     

Inhibition of Fibrinogen Reaction by Polysaccharide of Encapsulated Staphylococcus aureus

Encapsulated and nonencapsulated strains of Staphylococcus aureus which lack coagulase or clumping factor (bound coagulase), or both, were examined for the antigen associated with the fibrinogen-cell clumping reaction. Extracts of the cells were tested for the ability to react with fibrinogen or to inhibit fibrinogen precipitation. Antisera prepared against encapsulated (coagulase-positive, clumping factor-negative) variants, as well as against nonencapsulated wild-type (coagulase-positive, clumping factor-positive) S. aureus strains, contained high titers of clumping-inhibiting antibody. When coagulase-negative, clumping factor-negative mutants were the immunizing agents, antisera contained no demonstrable clumping-inhibiting antibody. Phenol extracts of all coagulase-positive strains tested precipitated fibrinogen, regardless of the ability of cells to clump in the presence of fibrinogen. Polysaccharide extracts of encapsulated, clumping factor-negative strains inhibited this fibrinogen-precipitating activity, whereas similar extracts of nonencapsulated staphylococci did not inhibit the fibrinogen reaction. From these results, it appeared that the coagulase-positive, encapsulated staphylococci which do not clump in fibrinogen solution possess clumping factor, but that their capsular polysaccharide inhibits clumping activity. These findings suggested a closer association of clumping factor and coagulase than is now recognized.     

A Comparison of Methods and the Validity of Deoxyribonuclease Tests for the Characterization of Staphylococci Isolated from Animals

Strains isolated from pigeons belonging to the coagulase-positive species Staphylococcus intermedius , coagulase-negative Staph. hyicus subsp. chromogenes strains from cattle and pigs, and Staph. aureus strains from poultry, gave weakly positive reactions in DNase plate culture tests and heat-resistant DNase tests. Staph. aureus and Staph. intermedius strains from other sources and coagulase-negative and coagulase-positive Staph. hyicus subsp. hyicus strains reacted strongly in these tests. A standardized plate culture test procedure is proposed and the use of DNase tests in the identification of staphylococci isolated from animals is discussed.