Lytic activities, protein profiles and morphologic characteristics of new bacteriophages isolated from canine and human Staphylococcus aureus strains.

The lytic activity, protein profile and morphology of five newly isolated phages from canine Staphylococcus aureus strains and one from a human S. aureus strain were compared with those of selected phages in the international phage sets (IPS). Five canine phages lysed 57 (76.0%) of 75 canine isolates of Staphylococcus aureus from Nigeria at routine test dilution (RTD) while 34 (IPS) phages typed only 31 (41.3%) strains at RTD or/and 100-RTD. The new human phage lysed 11 (14.7%) of 75 strains isolated from human diarrhoea. The new phages were readily propagated, specific in activity and stable during storage at 4 degrees C. Prominent proteins detected by SDS-PAGE indicated similarities between some of the phages but one canine phage was distinctly different, as was its morphology which was an isometric head with a short tail compared to oval heads and long tails which characterized others. IPS phages in the same serologic group had similar protein profiles but no correlation was observed with lytic groups. The use of protein profile and electron micrographs allowed classification of the phages into serogroups. It is concluded that the newly isolated canine phages could be very useful in typing Nigerian canine strains of S. aureus.     

Biochemical properties and differentiation of coagulase-positive staphylococci from rooks and gulls

Coagulase-positive staphylococci were found in the throats of 46 rooks (69 per cent) and 47 gulls (21 per cent) out of totals of 67 and 229 birds, respectively. Of 111 strains isolated from throat swabs 86 (77.5 per cent) were classified as Staphylococcus aureus and 25 strains (22.5 per cent) as Staphylococcus intermedius. Of the S aureus strains 82 per cent from rooks and 50 per cent from gulls were biotyped. Most of them were identified as biotypes D and B, only a few as biotype A, and none as biotype C. Moreover, among the 24 S intermedius strains (96 per cent) biotypes 1 and 2 could be differentiated according to the type of growth on crystal violet agar and their ability to produce acetoin and acid from maltose, mannitol and trehalose.     

Preliminary Observations on the Isolation of Escherichia coli Bacteriophages from the Intestinal Contents of Chickens

Eschericia coli bacteriophages were isolated from the intestines of chickens. These phages had different lytic patterns, and were propagated in nutrient broth containing 0.4 gm calcium chloride/litre. The agar layer technique was used to determine the routine test dilution (RTD) and plaque morphology. The phages differed in their 1) morphology, 2) RTD values, and their ability to lyse E. coli strains from various animals. All phages isolated lysed human K12 E. coli strains, whereas only two phages lysed the chicken E. coli strain. Phages isolated lysed E. coli from chicken, bovine, ovine, equine, and human, but not from porcine, canine and other avian species.     

Isolation and characteristics of bacteriophages from staphylococci of chicken origin.

Lysogenicity in chicken coagulase-positive staphylococci was tested by incubating the strains in the presence of mitomycin C. Of 88 strains tested, 84 (95.5%) were proved to be phage carriers and 81 were susceptible to any of the phages. The lysogenic strains were detected with almost equal frequency from both of typeable and untypeable strains by the international phages. Sixteen phages (CH phages) were isolated from chicken lysogenic strains, and their usefulness for the typing of chicken staphylococci was evaluated. Of 122 strains examined, 101 (82.8%) were found to be typeable with the CH phages at a routine test dilution (RTD). About 82% of strains untypeable by the phages of the international series were lysed by one or more of the CH phages. The phages seemed to be highly specific to chicken staphylococci, because they lysed only a few strains of animal origin other than chicken. Thus, the 16 phages newly established were found to have significant advantages in typing chicken strains.     

Bacteriophage typing of Staphylococcus hyicus subsp hyicus isolated from pigs

Four phages were isolated and used for typing Staphylococcus hyicus subsp hyicus isolated from pigs with or without exudative epidermatitis (EE) in Japan. Sixty-four (85.3%) of the 75 isolates examined were typeable at either routine test dilution (RTD) or 100 X RTD. Two or more kinds of phage patterns were present in the isolates from each pig with EE. All isolates from healthy pigs showed a single-phage pattern. Fourteen (32.6%) of 43 isolates and 7 (87.5%) of 8 isolates from pigs with EE in Belgium and Czechoslovakia, respectively, were typeable with the 4 phages. None of 180 isolates of S aureus, 7 (6.4%) of 110 isolates of S intermedius, and 2 (2.3%) of 86 isolates of S epidermidis were typeable.     

Staphylococcus aureus isolated from poultry in Australia. I. Phage typing and cultural characteristics

The phage typing and cultural characteristics of 574 strains of S. aureus of poultry origin in Australia were examined. With the avian phage set of Shimizu (1979) it was possible to type 74.2% of strains. A number of significant variations in the phage typing patterns of Australian strains compared to those reported from Japan and Europe were observed. A lower proportion of Australian strains were of avian phage group I and a higher proportion of group III. A high proportion of strains were of mixed lytic groups. No locally isolated phages were able to increase significantly the percentage of typeable strains, although four local phages appeared to be of greater value for phage typing poultry strains of S. aureus than some other phages of the avian phage set. The international (human) phage set was of limited value in typing Australian strains of poultry origin although four strains were identified which were indistinguishable from strains of human origin. Using cultural characteristics of the strains in conjunction with phage typing, the Australian strains of S. aureus were assigned to one of three major groups and nine subgroups. A list of typing phages considered to be valuable for use on Australian poultry strains of S. aureus is given.     

Species specificity in the adherence of staphylococci to canine and human corneocytes: a preliminary study

Staphylococcal pyoderma is rarely contagious between dogs and humans, or humans and dogs. This study investigated the hypothesis that there are species differences in adherence of Staphylococcus intermedius (the most common isolate from dogs) and Staphylococcus aureus (the most common isolate from humans) to canine and human corneocytes. Sheets of corneocytes were collected from the ventral abdomen of 10 dogs and the medial forearm of 10 humans (all healthy and without any history or physical signs of skin disease) using double-sided tape. Staphylococcus intermedius from a case of canine bacterial pyoderma and a human strain of S. aureus were prepared in phosphate buffered saline (PBS) and applied in duplicate, respectively, to the canine and human corneocyte-covered tapes using PBS as negative control. After incubation, rinsing, and staining with crystal violet, quantification of the adherent bacteria was carried out blindly by computerized image analysis. Staphylococcus intermedius was found to adhere significantly more to canine corneocytes than S. aureus (P = 0.0006), whereas S. aureus showed greater adherence to human corneocytes than S. intermedius (P < 0.0001). In addition, the pattern of adherence differed between the two organisms, with S. intermedius adhering to the entire surface and S. aureus adhering mainly to the periphery of both canine and human corneocytes. Preference for adherence to these two hosts may explain, in part, why S. intermedius and S. aureus are uncommonly isolated from human and canine skin infections, respectively.     

Phage typing of Staphylococcus aureus strains isolated in Sweden from bovine milk.

Both the human and the bovine international sets of phages were used for typing of 372 bovine Staphylococcus aureus (Sa) strains, whereas the bovine set alone was used for typing of a further 1183 strains. In addition, 338 of the strains were tested for antibiotic sensitivity. Out of 372 Sa strains 85.5% could be typed with the human and 89.8% with the bovine phage set. Of all the 1555 Sa strains used 92.4% were lysed by the bovine phage set. Several phage types can be present in one and the same herd and some of them can predominate. Resistance to most of the tested antibiotics was very low. The incidence of resistance to penicillin and ampicillin was 10.0% and 4.4% respectively.     

Characterization of Staphylococcus intermedius from pigeons, dogs, foxes, mink, and horses by pulsed-field gel electrophoresis

Staphylococcus intermedius from pigeons, dogs, foxes, mink, and horses, was characterized by pulsed-field gel electrophoresis (PFGE) to evaluate the use of this typing method for discriminating among strains. SmaI cut the chromosomal DNA into 7-13 fragments ranging from approximately 48 kb to 655 kb, with most of the detectable fragments being smaller than 172 kb. S. intermedius from various animals had a high degree of restriction fragment length polymorphism. Pigeon strains have a similar genotype, despite the difference in their isolation area. Phage typing indicated that the dog, fox, and mink strains belong to the canine I or canine II type. The PFGE method further differentiated the mink strains from the dog and fox strains with regard to three fragments between 256 kb and 570 kb. As such, genomic DNA fingerprinting by PFGE appears to be an effective technique for discriminating S. intermedius strains from various animals. A combination of PFGE typing and phage typing would provide more detailed information than the single method for ecological investigations of S. intermedius.     

Characteristics of Staphylococcus aureus strains from pathological processes in swine

120 Staphylococcus aureus strains from different and sporadic pathological process in swines have been investigated on biochemical properties and phage patterns. 98 strains (82%) exhibit coagulation of human plasma, no coagulation of bovine plasma, crystal-violet-type C, hemolysin-type beta, no formation of fibrinolysine and sensitivity for phages of group I, III and M. Only in few strains we observed reactions with phages of group II. 26 from 66 strains were resistant to antibiotics, and among them 24 showed resistance to oxytetracacline.     

The use of a new phage set for typing poultry strains of Staphylococcus aureus obtained from seven countries

1. Eighty per cent of poultry strains of Staphylococcus aureus tested from French, Belgian, English, West German, Japanese and Argentinian sources were typable using a set of typing phages isolated in this laboratory. Strains from Bulgaria, however, with few exceptions were not typable with this phage set. 2. Strains isolated from lesions generally resembled those from apparently healthy poultry. 3. The existence of two distinct Staph. aureus biotypes on poultry was confirmed by isolates from six of the countries; one of these biotypes closely resembled Staph. aureus variety gallinae as described by Witte et al. (1977).     

Establishment of a new bacteriophage set for typing avian staphylococci.

A new phage set was proposed for the typing of coagulase-positive staphylococci from chickens. The set was composed of 16 phages isolated from lysogenic staphylococci from chickens and 3 adapted phages derived from the international phage set. These phages were classified into 4 groups (I, II, III and IV) according to their host ranges. The 19 phages were used for typing 569 coagulase-positive staphylococci; 94.2% of the isolates were lysed by 1 or more phages, 88.0% at a routine test dilution, and 6.25% only at a routine test dilution X 100. The reacting ability of the phages seemed to be limited to staphylococci from chickens. It was proved that the phages were more accurate than the international phage set in typing staphylococci isolated from chicken in Japan.     

Staphylococcal colonization of mucosal and lesional skin sites in atopic and healthy dogs

Staphylococcal colonization was compared in healthy dogs and in dogs with atopic dermatitis. Bacterial swabs were collected from the nasal mucosa, ear and perineum of 43 healthy and 24 atopic dogs and also from potentially infected skin lesions of the atopic dogs. Coagulase positive staphylococcal isolates were identified to the species level. At the time of this study Staphylococcus intermedius was considered a single species but has since been recognized as comprising at least three species with canine isolates believed to belong to Staphylococcus pseudintermedius . Of atopic dogs, 87.5% were colonized with S. intermedius compared to only 37.2% of healthy dogs. The ear was the only carriage site that showed any significant difference in S. intermedius isolation between healthy and atopic dogs. The perineum represented the most frequently colonized mucosal site for both groups. Sampling the nasal mucosa alone identified 71.4% of atopic and 37.5% of healthy S. intermedius carriers. Inclusion of a perineal swab identified 100% of atopic and 93.8% of healthy carriers. S. intermedius was isolated from all the lesional sites sampled from atopic dogs. Significantly fewer dogs were colonized by Staphylococcus aureus than S. intermedius , and there was no significant difference between S. aureus colonization of atopic and healthy dogs. S. aureus was not recovered from any lesions in atopic dogs. The results show that S. intermedius carriage is more prevalent in atopic dogs compared to healthy dogs and that to identify staphylococcal carriers both the nasal mucosa and the perineum should be sampled.     

Identification of coagulase-positive staphylococci isolated from bovine milk

A total of 414 coagulase-positive staphylococcal strains obtained at the mastitis laboratory, National Veterinary Institute, Uppsala, Sweden, were studied. One hundred and seventy seven strains were used for a frequency study. Ninety-seven per cent were identified as Staphylococcus aureus, 2% as Staphylococcus intermedius and 1% as Staphylococcus hyicus. Two hundred and thirty seven strains with atypical hemolysis reactions on bovine blood agar were randomly selected, with the aim to increase the number of S. intermedius and S. hyicus strains available for testing. Eight different characteristics, including physiological, enzymatical and biochemical properties, were used to identify the coagulase-positive Staphylococcus species. The results of this study suggest that the following tests should be included for correct identification of the 3 different species of coagulase-positive staphylococci: P agar supplemented with acriflavin, beta-galactosidase and hemolytic reaction on chocolate agar. These 3 tests are simple and quick to perform and enable accurate for easy differentiation of the 3 coagulase-positive Staphylococcus species.     

Characteristics of Staphylococcus aureus isolated from lesions of horses.

Seventy-six Staphylococcus aureus strains isolated from various lesions of horses were characterized. All of the 76 strains were identified as biotypes B (38.2%) and C (61.8%). Of 55 strains tested, 42 (76.4%) were differentiated into 7 coagulase types. Coagulase types V and VII were predominant in the metritis strains. Coagulase type II was found most frequently in the strains from phlegmon, dermatitis, sinusitis, empyema sinus, and nasal catarrh. Forty-two (55.3%) of the 76 strains were differentiated into 24 phage patterns. Twenty (58.8%) of 34 typable strains from metritis were lysed by the human group I phage 52, and group II phages 3A, 3C, 55 and 71. Forty-five (59.2%) of the 76 strains were resistant to 1 or more of 6 antibiotics. Strains resistant to penicillin G, irrespective of source, were most frequent (95.6%). Forty (93.0%) of 43 strains resistant to penicillin G alone or in combination with other antibiotics produced beta-lactamase. Only 8 (10.5%) of the 76 strains produced enterotoxins A (n = 2), B (n = 1) or C (n = 5), and they all were isolated from metritis. Only 1 strain isolated from phlegmon and 2 from metritis produced exfoliative toxin (ET) and toxic shock syndrome toxin-1 (TSST-1), respectively. The latter 2 strains also produced enterotoxin C. The results of the present study showed the first evidence of the presence of both ET- and TSST-1-producing S. aureus isolated from horses.     

Microbiological features of Staphylococcus schleiferi subsp. coagulans, isolated from dogs and possible misidentification with other canine coagulase-positive staphylococci

Staphylococcus schleiferi subsp. coagulans has only rarely been isolated and identified from the external auditory meatus of dogs suffering from external otitis. Its morphological and basic biochemical characteristics are of relatively little value for identification, as it phenotypically resembles another coagulase-positive staphylococci (CPS) and, consequently, may be easily misidentified as S. intermedius or even as S. aureus. In the present work, differentiation of S. schleiferi ssp. coagulans was therefore based on specific biochemical and genetic methods. All the strains were evaluated with the following commercial methods: Api Staph System (bioMérieux, Marcy l'Etoil, France), BBL Crystal Identification Systems (Gram-Positive ID Kit and Rapid Gram-Positive ID Kit; Becton Dickinson), and GEN-PROBE AccuProbe, Staphylococcus aureus identification test (bioMérieux). Gram-Positive ID System/GP database includes the broadest range of staphylococcal species and correctly identifies the majority of strains important in veterinary medicine. Therefore, it is an acceptable alternative to conventional methods for identification of canine staphylococcal isolates. Reliable differentiation of S. aureus from S. schleiferi ssp. coagulans and S. intermedius was feasible with AccuProbe for S. aureus, which gave positive results only for S. aureus; all other CPS tested were negative.     

What’s happened to Staphylococcus intermedius? Taxonomic revision and emergence of multi‐drug resistance

Staphylococcus intermedius has been the predominant coagulase-positive Staphylococcus isolated from canine skin and mucosae and the most commonly reported staphylococcal pathogen in small animal practice for the last 35 years. Although microbiological tests have historically indicated variability in biochemical characteristics amongst S. intermedius isolates from animals, an acceptable level of diagnostic accuracy for clinical purposes was readily achievable with routine phenotypic testing. However, three recent developments have changed our understanding of the term "S. intermedius" and have challenged veterinary bacteriologists to ensure correct species identification of pathogenic staphylococci from small animals. First, the increasing recognition of meticillin-resistant Staphylococcus aureus in small animal practice and its human health implications demand accurate species identification. Secondly, the application of molecular techniques to analysis of staphylococcal isolates has led to a revised taxonomy and canine isolates of S. intermedius being re-named S. pseudintermedius. Thirdly, the recent, rapid emergence of meticillin- and multi-drug-resistant strains of Staphylococcus pseudintermedius (MRSP) has become a major therapeutic challenge in veterinary practice worldwide, including the UK. This article discusses the background of the recent taxonomic changes within the genus Staphylococcus and reviews the key features of MRSP and its implications for day-to-day laboratory diagnosis and small animal practice.     

In vitro antimicrobial activity of orbifloxacin against Staphylococcus intermedius isolates from canine skin and ear infections

The objective of the study was to evaluate the in vitro activity of orbifloxacin against Staphylococcus intermedius strains isolated in France from canine skin and ear infections. The minimum inhibitory concentrations (MICs) of orbifloxacin against 240 field S. intermedius isolates (69 skin and 171 ear isolates) ranged from 0.016 to 8 mg l(-1), with MIC50 and MIC90 equal to 0.5 and 1 mg l(-1), respectively. Only one strain, a pyoderma isolate was resistant (MIC=8 mg l(-1)). Orbifloxacin was tested at different concentrations for killing rate against five isolates obtained from pyoderma cases and against a reference strain (Staphylococcus aureus ATCC 29213). Orbifloxacin expressed a concentration-dependent bactericidal activity against the S. aureus reference strain, but a time-dependent bactericidal activity against S. intermedius. Orbifloxacin induced bactericidal effect against the S. intermedius strains tested with concentrations equal to or two times MIC.     

Characteristics of Staphylococcus aureus from subclinical bovine mastitis in Brazil

A total of 127 Staphylococcus aureus strains isolated from milk samples of cows with subclinical bovine mastitis was examined for biotype, phage pattern, in-vitro antibiotic susceptibilities and ability to produce enterotoxins. The majority of the strains showed features consistent with bovine rather than human origin. All strains were sensitive to the antibiotics tested, except penicillin and streptomycin. Enterotoxigenicity was observed in 6 (4.7%) strains and only enterotoxins A and C were produced.     

A comparison of adherence by four strains of Staphylococcus intermedius and Staphylococcus hominis to canine corneocytes collected from normal dogs and dogs suffering from atopic dermatitis

The aim of this study was to compare the adherence of four strains of Staphylococcus intermedius and a single strain of Staphylococcus hominis to corneocytes from both normal dogs and dogs suffering from atopic dermatitis. Cells from the skin surface, corneocytes, were collected from 10 normal dogs and 10 dogs suffering from atopic dermatitis. Four strains of S. intermedius, three isolated from canine pyoderma skin lesions (strains A, B and C), and one isolated form from canine synovial membrane sample from a case of septic arthritis (strain D) were compared. S. hominis, which is not normally associated with canine disease, was also evaluated for its ability to adhere to canine corneocytes. S. hominis did not adhere to canine corneocytes. All four strains of S. intermedius adhered well to canine corneocytes collected from both normal and atopic dogs. All strains of S. intermedius showed statistically greater adherence to corneocytes collected from atopic dogs compared with those collected from normal dogs. It was concluded that the adherence assay employed here showed that S. hominis does not adhere to canine corneocytes, S. intermedius adheres preferentially to atopic corneocytes.