Vol. 59, No. 8
INFECTION AND IMMUNITY, Aug. 1991, p. 2615-2623
0019-9567/91/082615-09$02.00/0 Copyright C) 1991, American Society for Microbiology
Experimental Staphylococcus TOMAS
aureus
Arthritis in Mice
BREMELL,1* STEFAN LANGE,' ALIA YACOUB,2 CECILIA RYDEN,2'3
AND
ANDRZEJ TARKOWSKI'
Departments of Rheumatology and Clinical Bacteriology, Immunology and Virology, University of Goteborg, S-413 46 Goteborg,' and Departments of Medical and Physiological Chemistry2 and Infectious Diseases,3 University of Uppsala, Uppsala, Sweden Received 28 January 1991/Accepted 15 May 1991
Staphylococcus aureus arthritis is usually caused by bacteremia and is highly destructive. Controlled studies septic arthritis in humans are difficult to perform, because the time of onset of the infection is unknown. Animal models of bacterial arthritis make it possible to control important variables in experimental studies. We present a mouse model of S. aureus arthritis in which the intravenous administration of 107 cells of S. aureus LS-1 induced arthritis or osteitis or both within 3 weeks in 80 to 90% of the mice. Signs of arthritis emerged within the first few days after the injection. An interesting finding was that the S. aureus strain used in this study binds bone sialoprotein, a glycoprotein known to be specifically localized to bone tissue. This new model of S. aureus arthritis enables the study of the kinetics of joint destruction and the host-bacterium relationship as well as therapeutical approaches to septic arthritis and osteomyelitis. on
Bacterial arthritis is a rapidly progressive and highly destructive joint disease in humans. All destructive joint diseases, including inflammatory disorders such as rheumatoid arthritis, are connected to an increased incidence of bacterial arthritis (6). Certain forms of therapy such as joint implants and immunosuppressive treatment display an increased frequency of bacterial arthritis (6). Staphylococcus aureus is the causative agent in about 60% of nongonococcal bacterial arthritis cases (4). In patients with rheumatic diseases, this value is even higher, approaching 75% (3). Laboratory models of bacterial arthritis have been used previously (5, 12, 16, 21, 24, 25, 27-29). In most instances bacteria have been injected intraarticularly, usually utilizing the rabbit as an experimental animal. In contrast, human bacterial arthritis usually has a hematogenic spread (6). We have recently reported a spontaneous outbreak of S. aureus arthritis in our mouse colony (1). In this report, we describe a murine model of bacterial arthritis following injection of an S. aureus strain, LS-1, originating from a spontaneously arthritic mouse. Another S. aureus strain, 024, originating from a patient with acute osteomyelitis, has also been used. Furthermore, binding of bone sialoprotein, a glycoprotein of the extracellular matrix of bone tissue, fibronectin, and collagen type I, to the S. aureus strains is analyzed.
Bacterial strains and culture conditions. S. aureus LS-1 originally isolated from a swollen joint of a spontaneously arthritic NZB/W mouse (1). This bacterial strain proved to be catalase and coagulase positive and displayed the following phage type: 6147153154175177/85188+. Before each experiment, bacteria were cultured on blood agar (5% human erythrocytes) for 24 h and then reincubated on blood agar for another 24 h. A bacterial solution was prepared, using McFarland nephelometer no. 8 (7), and then further diluted in physiological saline to the desired concentration and used live or formalin killed as indicated below. S. aureus 024, isolated from a patient suffering from osteomyelitis (23), and S. epidermidis 7686 from a patient with a peritoneal catheter, a kind gift from G. Pulverer, University of Cologne, Cologne, Federal Republic of Germany, were both used as control strains in the protein binding assays. S. aureus 024 was also used in the study. Administration of bacteria. Intravenous (i.v.) injections of 0.2 ml of S. aureus suspension in physiological saline were given in the tail. Control mice were injected with 0.2 ml of either formalin-treated (4%, 20 min) S. aureus or physiological saline. Evaluation of arthritis and osteitis. All mice were followed individually. Limbs were inspected visually by two observers (T.B. and A.T.) at regular intervals (6 h and 1, 2, 3, 7, 10, 14, 21, and 28 days after inoculation). Arthritis was defined as visible joint swelling or erythema of at least one joint or both. The mean number of arthritic limbs per animal was registered. Nodose tail was reported when there was visible and palpable localized swelling of the tail. The overall condition was evaluated, assessing weight, general appearance, alertness, and skin abnormalities of each mouse. Experimental protocol. (i) Susceptibility to arthritis of different mouse strains. Fifteen Swiss, 14 BALB/c, 17 C3H/ HeJ, 16 C3H/HeN, and 14 C57BL mice were used. All mice were 8 weeks old. Each mouse strain was divided into three groups: controls receiving physiological saline and probands challenged with 107 or 108 S. aureus LS-1. During a 5-week period, the mice were evaluated with regard to joint swelling, nodose tail, and overall condition. was
MATERIALS AND METHODS Mice. Swiss and BALB/c (original breeding stock; ALAB, Stockholm, Sweden), C3H/HeJ (original breeding stock; Jackson Laboratories, Bar Harbor, Maine), C3H/HeN (original breeding stock; Charles River Laboratories, Margate, Kent, United Kingdom), and C57BL (original breeding stock; Bomholtgard, Ry, Denmark) mice were bred in the animal facility at the Department of Clinical Bacteriology, Immunology and Virology, University of Goteborg, Goteborg, Sweden. Only male mice were used. They were fed laboratory chow and water ad libitum under standard conditions of temperature and light. *
Corresponding author. 2615
2616
INFECT. IMMUN.
BREMELL ET AL.
TABLE 1. Occurrence of arthritis and nodose tail at sacrifice in 13-week-old mice of five different strains challenged 5 weeks earlier by i.v. injection with S. aureus LS-1 Mouse strain
Swiss
BALB/c
C3H/HeN
C57BL
No. of mice
No. of
arthritic
mice
with nodose tail
mice that died
107 108
5 5 5
0 3 1
0 0 0
0 1 1
0 107
4 5 5
0 0 1
0 0 0
0 0 2
107 108
5 6 6
0 1 2
0 3 2
0 1 0
0 107
6 5
108
5
0 0 1
0 1 1
0 1 3
0
4 5 5
0 0 0
0 1 0
0 0 2
0
108 C3H/HeJ
No. of
No. of mice
Dose
(bacteria)
0
io7 108
(ii) Influence of dose of S. aureus and route of administration on arthritis induction. Sixty-four 3- to 6-week-old Swiss mice were administered six different doses of S. aiureus LS-1 (107, 2.5 x 106, 2.5 x 105, 2.5 x 104, 2.5 x 103, 2.5 x 102, and 0 bacteria per mouse) i.v., intraperitoneally, and subcutaneously. At sacrifice 5 weeks later, bacterial isolates were obtained from blood, joints, and tail of eight selected mice (two of which were controls). (iii) Kinetics of arthritis and bacterial growth. Eighty 3- to 6-week-old Swiss mice were each given 107 S. aureus LS-1 i.v., and 15 control mice were injected i.v. with physiological saline. Another 10 mice received 107 killed S. aureus bacteria. Groups of 10 mice injected with live bacteria were sacrificed at selected intervals, i.e., after 6 h, 24 h, 48 h, 72 h, 7 days, 14 days, 21 days, and 28 days. The 15 saline control mice were sacrificed after 24 h (5 mice), 7 days (5 mice), and 28 days (5 mice). The 10 control mice injected with killed bacteria were sacrificed after 28 days. After sacrifice, both the talocrural and the radiocarpal joints, as well as the tails, were dissected, and bacterial samples were obtained. Blood samples were also obtained from all animals. The samples were transferred to agar containing 5% human blood and incubated for 48 h at 37°C. Bacterial colonies were tested for catalase and coagulase activity. To avoid false-positive results due to contamination, a joint, tail, or blood sample was considered positive when more than 20 colonies of S. aureus were present. Fourteen S. aureus isolates were also tested for bacteriophage pattern. In four mice injected with live bacteria, isolates were taken from liver, spleen, heart, lungs, and kidneys on day 28. The mice were weighed on one to four occasions during the experiment. Ten 4-week-old Swiss mice were injected i.v. with 107 S. aureus 024. They were regularly evaluated during 5 weeks with regard to joint swelling, nodose tail, and overall condi-
tion. Histopathologic examination. To evaluate the histopatho-
logic features of the joint disease, a limited number of mice inoculated i.v. with 107 S. aureus were sacrificed on days 3 and 14 after inoculation. Histologic examination was performed after routine fixation, decalcification, paraffin embedding, and staining with hematoxylin and eosin. Binding of radiolabeled proteins to staphylococcal cells. Bone sialoprotein purified from bovine bone (2) was kindly provided by Dick Heinegard, Department of Medical and Physiological Chemistry, University of Lund, Lund, Sweden, and was radiolabeled to a specific activity of 2 x 107 cpm/pLg with 1251 (Radiochemical Centre, Amersham, United Kingdom), using the lodobeads (Pierce Chemical Co., Rockford, Il.) method (17). Fibronectin was purified from human plasma (19), and collagen type I from calf skin (Vitrogen 100) was obtained from the Collagen Corp., Palo Alto, Calif. These proteins were radiolabeled by the above-mentioned method, yielding specific activities of 2 x 107 cpm/pLg for fibronectin and 107 cpm/lLg for collagen type I. Bacteria grown on blood agar were transferred to tryptic soy broth (Oxoid Ltd., Basingstock, United Kingdom) and cultured at 37°C for 16 h. Prior to experiments, bacteria were washed twice with cold phosphate-buffered saline (0.13 M NaCl, 10 mM sodium phosphate, pH 7.4) and subsequently resuspended in a buffer containing 137 mM NaCl, 5 mM KCI, 0.7 mM MgSO4, 1.2 mM CaCl, 10 mM HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid), and 100
80
(.) t: 60
0L)C.) E 0
40
20
10
0
20
30
Days after inoculation No. of mice
80
60
70
39
48
10
28
29
19
FIG. 1. Prevalence of arthritis in Swiss mice injected i.v. with 107 S. aureus. Note that the number of animals decreases with time due to sacrifice.
EXPERIMENTAL S. AUREUS ARTHRITIS
VOL. 59, 1991
a
2617
b
_^ __
.,
._ \ .w
.
FIG. 2. Photograph showing a normal (a) and an arthritic (b) hindpaw of an 8-week-old male Swiss mouse 28 days after i.v. injection with 107 S. aureus LS-1.
0.1% ovalbumin, pH 7.4 (buffer A), at a concentration of 1010 bacteria per ml. The binding of staphylococcal cells to bone sialoprotein was performed as described previously (23). The binding of collagen type I was performed with denatured collagen (56°C for 20 min). The denaturation was performed prior to radiolabeling and then again just before use in the binding assay. The experiments with collagen binding were performed at room temperature to minimize renaturation of the collagen during the binding procedure. Briefly, 10 ng of 1251I-labeled protein was incubated with 109 bacteria in 500 ,ul of buffer A. Incubations were performed end-over-end at 4°C for bone sialoprotein and fibronectin and at 22°C for collagen type I. Binding was detected at defined intervals. A 100-,ul portion of bacterial suspension was layered on a Percoll gradient and centrifuged, and the supernatant was discarded. Radioactivity in the pellet was measured in a gamma counter. Values are means of double samples, and the time curve was performed three times. In all experiments a positive control, S. aureus 024, as well as a negative control strain, S. epidermidis 7686, were used. Statistics. Comparisons regarding weights (see Table 2) were made by Student's two-tailed t test. All values reported are means + standard errors, if not stated otherwise. RESULTS Establishment of the model of S. aureus arthritis. Five different mouse strains, Swiss, BALB/c, C3H/HeJ, C3H/
HeN, and C57BL, were used in experiment A to study the genetic susceptibility of development of arthritis. Swiss mice displayed the highest occurrence of arthritis (Table 1), and in all subsequent experiments only this mouse strain was used. To establish the optimal infectious dose of bacteria and the
4
FIG. 3. Photograph showing arthritis of the right forepaw 28 days after i.v. injection with 107 S. aureus LS-1. Note the nonarthritic left forepaw.
2618
INFECT. IMMUN.
BREMELL ET AL. 2.0
1.5
0
10
.0
6 0.5
0
0
10
20
30
Days after inoculation 39 28 eeeeeeeeeeeeeeeeeeeeeeeeeeeeee10 xo60 No. of 19 29 mice XXX7048 FIG. 4. Occurrence of infectious arthritis expressed as the mean number of arthritic limbs per mouse +standard error after a single i.v. injection of 107 S. aureus LS-1. The number of animals decreases with time due to sacrifice.
best route of administration, 64 mice were studied in experiment B. The mice injected i.v. displayed the highest proportion of arthritis, especially those given 107 bacteria. Thus, 75% of the mice injected with 107 S. aureus cells displayed arthritis and nodose tails. In contrast, when given
INFECTION AND IMMUNITY, Aug. 1991, p. 2615-2623
0019-9567/91/082615-09$02.00/0 Copyright C) 1991, American Society for Microbiology
Experimental Staphylococcus TOMAS
aureus
Arthritis in Mice
BREMELL,1* STEFAN LANGE,' ALIA YACOUB,2 CECILIA RYDEN,2'3
AND
ANDRZEJ TARKOWSKI'
Departments of Rheumatology and Clinical Bacteriology, Immunology and Virology, University of Goteborg, S-413 46 Goteborg,' and Departments of Medical and Physiological Chemistry2 and Infectious Diseases,3 University of Uppsala, Uppsala, Sweden Received 28 January 1991/Accepted 15 May 1991
Staphylococcus aureus arthritis is usually caused by bacteremia and is highly destructive. Controlled studies septic arthritis in humans are difficult to perform, because the time of onset of the infection is unknown. Animal models of bacterial arthritis make it possible to control important variables in experimental studies. We present a mouse model of S. aureus arthritis in which the intravenous administration of 107 cells of S. aureus LS-1 induced arthritis or osteitis or both within 3 weeks in 80 to 90% of the mice. Signs of arthritis emerged within the first few days after the injection. An interesting finding was that the S. aureus strain used in this study binds bone sialoprotein, a glycoprotein known to be specifically localized to bone tissue. This new model of S. aureus arthritis enables the study of the kinetics of joint destruction and the host-bacterium relationship as well as therapeutical approaches to septic arthritis and osteomyelitis. on
Bacterial arthritis is a rapidly progressive and highly destructive joint disease in humans. All destructive joint diseases, including inflammatory disorders such as rheumatoid arthritis, are connected to an increased incidence of bacterial arthritis (6). Certain forms of therapy such as joint implants and immunosuppressive treatment display an increased frequency of bacterial arthritis (6). Staphylococcus aureus is the causative agent in about 60% of nongonococcal bacterial arthritis cases (4). In patients with rheumatic diseases, this value is even higher, approaching 75% (3). Laboratory models of bacterial arthritis have been used previously (5, 12, 16, 21, 24, 25, 27-29). In most instances bacteria have been injected intraarticularly, usually utilizing the rabbit as an experimental animal. In contrast, human bacterial arthritis usually has a hematogenic spread (6). We have recently reported a spontaneous outbreak of S. aureus arthritis in our mouse colony (1). In this report, we describe a murine model of bacterial arthritis following injection of an S. aureus strain, LS-1, originating from a spontaneously arthritic mouse. Another S. aureus strain, 024, originating from a patient with acute osteomyelitis, has also been used. Furthermore, binding of bone sialoprotein, a glycoprotein of the extracellular matrix of bone tissue, fibronectin, and collagen type I, to the S. aureus strains is analyzed.
Bacterial strains and culture conditions. S. aureus LS-1 originally isolated from a swollen joint of a spontaneously arthritic NZB/W mouse (1). This bacterial strain proved to be catalase and coagulase positive and displayed the following phage type: 6147153154175177/85188+. Before each experiment, bacteria were cultured on blood agar (5% human erythrocytes) for 24 h and then reincubated on blood agar for another 24 h. A bacterial solution was prepared, using McFarland nephelometer no. 8 (7), and then further diluted in physiological saline to the desired concentration and used live or formalin killed as indicated below. S. aureus 024, isolated from a patient suffering from osteomyelitis (23), and S. epidermidis 7686 from a patient with a peritoneal catheter, a kind gift from G. Pulverer, University of Cologne, Cologne, Federal Republic of Germany, were both used as control strains in the protein binding assays. S. aureus 024 was also used in the study. Administration of bacteria. Intravenous (i.v.) injections of 0.2 ml of S. aureus suspension in physiological saline were given in the tail. Control mice were injected with 0.2 ml of either formalin-treated (4%, 20 min) S. aureus or physiological saline. Evaluation of arthritis and osteitis. All mice were followed individually. Limbs were inspected visually by two observers (T.B. and A.T.) at regular intervals (6 h and 1, 2, 3, 7, 10, 14, 21, and 28 days after inoculation). Arthritis was defined as visible joint swelling or erythema of at least one joint or both. The mean number of arthritic limbs per animal was registered. Nodose tail was reported when there was visible and palpable localized swelling of the tail. The overall condition was evaluated, assessing weight, general appearance, alertness, and skin abnormalities of each mouse. Experimental protocol. (i) Susceptibility to arthritis of different mouse strains. Fifteen Swiss, 14 BALB/c, 17 C3H/ HeJ, 16 C3H/HeN, and 14 C57BL mice were used. All mice were 8 weeks old. Each mouse strain was divided into three groups: controls receiving physiological saline and probands challenged with 107 or 108 S. aureus LS-1. During a 5-week period, the mice were evaluated with regard to joint swelling, nodose tail, and overall condition. was
MATERIALS AND METHODS Mice. Swiss and BALB/c (original breeding stock; ALAB, Stockholm, Sweden), C3H/HeJ (original breeding stock; Jackson Laboratories, Bar Harbor, Maine), C3H/HeN (original breeding stock; Charles River Laboratories, Margate, Kent, United Kingdom), and C57BL (original breeding stock; Bomholtgard, Ry, Denmark) mice were bred in the animal facility at the Department of Clinical Bacteriology, Immunology and Virology, University of Goteborg, Goteborg, Sweden. Only male mice were used. They were fed laboratory chow and water ad libitum under standard conditions of temperature and light. *
Corresponding author. 2615
2616
INFECT. IMMUN.
BREMELL ET AL.
TABLE 1. Occurrence of arthritis and nodose tail at sacrifice in 13-week-old mice of five different strains challenged 5 weeks earlier by i.v. injection with S. aureus LS-1 Mouse strain
Swiss
BALB/c
C3H/HeN
C57BL
No. of mice
No. of
arthritic
mice
with nodose tail
mice that died
107 108
5 5 5
0 3 1
0 0 0
0 1 1
0 107
4 5 5
0 0 1
0 0 0
0 0 2
107 108
5 6 6
0 1 2
0 3 2
0 1 0
0 107
6 5
108
5
0 0 1
0 1 1
0 1 3
0
4 5 5
0 0 0
0 1 0
0 0 2
0
108 C3H/HeJ
No. of
No. of mice
Dose
(bacteria)
0
io7 108
(ii) Influence of dose of S. aureus and route of administration on arthritis induction. Sixty-four 3- to 6-week-old Swiss mice were administered six different doses of S. aiureus LS-1 (107, 2.5 x 106, 2.5 x 105, 2.5 x 104, 2.5 x 103, 2.5 x 102, and 0 bacteria per mouse) i.v., intraperitoneally, and subcutaneously. At sacrifice 5 weeks later, bacterial isolates were obtained from blood, joints, and tail of eight selected mice (two of which were controls). (iii) Kinetics of arthritis and bacterial growth. Eighty 3- to 6-week-old Swiss mice were each given 107 S. aureus LS-1 i.v., and 15 control mice were injected i.v. with physiological saline. Another 10 mice received 107 killed S. aureus bacteria. Groups of 10 mice injected with live bacteria were sacrificed at selected intervals, i.e., after 6 h, 24 h, 48 h, 72 h, 7 days, 14 days, 21 days, and 28 days. The 15 saline control mice were sacrificed after 24 h (5 mice), 7 days (5 mice), and 28 days (5 mice). The 10 control mice injected with killed bacteria were sacrificed after 28 days. After sacrifice, both the talocrural and the radiocarpal joints, as well as the tails, were dissected, and bacterial samples were obtained. Blood samples were also obtained from all animals. The samples were transferred to agar containing 5% human blood and incubated for 48 h at 37°C. Bacterial colonies were tested for catalase and coagulase activity. To avoid false-positive results due to contamination, a joint, tail, or blood sample was considered positive when more than 20 colonies of S. aureus were present. Fourteen S. aureus isolates were also tested for bacteriophage pattern. In four mice injected with live bacteria, isolates were taken from liver, spleen, heart, lungs, and kidneys on day 28. The mice were weighed on one to four occasions during the experiment. Ten 4-week-old Swiss mice were injected i.v. with 107 S. aureus 024. They were regularly evaluated during 5 weeks with regard to joint swelling, nodose tail, and overall condi-
tion. Histopathologic examination. To evaluate the histopatho-
logic features of the joint disease, a limited number of mice inoculated i.v. with 107 S. aureus were sacrificed on days 3 and 14 after inoculation. Histologic examination was performed after routine fixation, decalcification, paraffin embedding, and staining with hematoxylin and eosin. Binding of radiolabeled proteins to staphylococcal cells. Bone sialoprotein purified from bovine bone (2) was kindly provided by Dick Heinegard, Department of Medical and Physiological Chemistry, University of Lund, Lund, Sweden, and was radiolabeled to a specific activity of 2 x 107 cpm/pLg with 1251 (Radiochemical Centre, Amersham, United Kingdom), using the lodobeads (Pierce Chemical Co., Rockford, Il.) method (17). Fibronectin was purified from human plasma (19), and collagen type I from calf skin (Vitrogen 100) was obtained from the Collagen Corp., Palo Alto, Calif. These proteins were radiolabeled by the above-mentioned method, yielding specific activities of 2 x 107 cpm/pLg for fibronectin and 107 cpm/lLg for collagen type I. Bacteria grown on blood agar were transferred to tryptic soy broth (Oxoid Ltd., Basingstock, United Kingdom) and cultured at 37°C for 16 h. Prior to experiments, bacteria were washed twice with cold phosphate-buffered saline (0.13 M NaCl, 10 mM sodium phosphate, pH 7.4) and subsequently resuspended in a buffer containing 137 mM NaCl, 5 mM KCI, 0.7 mM MgSO4, 1.2 mM CaCl, 10 mM HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid), and 100
80
(.) t: 60
0L)C.) E 0
40
20
10
0
20
30
Days after inoculation No. of mice
80
60
70
39
48
10
28
29
19
FIG. 1. Prevalence of arthritis in Swiss mice injected i.v. with 107 S. aureus. Note that the number of animals decreases with time due to sacrifice.
EXPERIMENTAL S. AUREUS ARTHRITIS
VOL. 59, 1991
a
2617
b
_^ __
.,
._ \ .w
.
FIG. 2. Photograph showing a normal (a) and an arthritic (b) hindpaw of an 8-week-old male Swiss mouse 28 days after i.v. injection with 107 S. aureus LS-1.
0.1% ovalbumin, pH 7.4 (buffer A), at a concentration of 1010 bacteria per ml. The binding of staphylococcal cells to bone sialoprotein was performed as described previously (23). The binding of collagen type I was performed with denatured collagen (56°C for 20 min). The denaturation was performed prior to radiolabeling and then again just before use in the binding assay. The experiments with collagen binding were performed at room temperature to minimize renaturation of the collagen during the binding procedure. Briefly, 10 ng of 1251I-labeled protein was incubated with 109 bacteria in 500 ,ul of buffer A. Incubations were performed end-over-end at 4°C for bone sialoprotein and fibronectin and at 22°C for collagen type I. Binding was detected at defined intervals. A 100-,ul portion of bacterial suspension was layered on a Percoll gradient and centrifuged, and the supernatant was discarded. Radioactivity in the pellet was measured in a gamma counter. Values are means of double samples, and the time curve was performed three times. In all experiments a positive control, S. aureus 024, as well as a negative control strain, S. epidermidis 7686, were used. Statistics. Comparisons regarding weights (see Table 2) were made by Student's two-tailed t test. All values reported are means + standard errors, if not stated otherwise. RESULTS Establishment of the model of S. aureus arthritis. Five different mouse strains, Swiss, BALB/c, C3H/HeJ, C3H/
HeN, and C57BL, were used in experiment A to study the genetic susceptibility of development of arthritis. Swiss mice displayed the highest occurrence of arthritis (Table 1), and in all subsequent experiments only this mouse strain was used. To establish the optimal infectious dose of bacteria and the
4
FIG. 3. Photograph showing arthritis of the right forepaw 28 days after i.v. injection with 107 S. aureus LS-1. Note the nonarthritic left forepaw.
2618
INFECT. IMMUN.
BREMELL ET AL. 2.0
1.5
0
10
.0
6 0.5
0
0
10
20
30
Days after inoculation 39 28 eeeeeeeeeeeeeeeeeeeeeeeeeeeeee10 xo60 No. of 19 29 mice XXX7048 FIG. 4. Occurrence of infectious arthritis expressed as the mean number of arthritic limbs per mouse +standard error after a single i.v. injection of 107 S. aureus LS-1. The number of animals decreases with time due to sacrifice.
best route of administration, 64 mice were studied in experiment B. The mice injected i.v. displayed the highest proportion of arthritis, especially those given 107 bacteria. Thus, 75% of the mice injected with 107 S. aureus cells displayed arthritis and nodose tails. In contrast, when given
