JCM Accepted Manuscript Posted Online 15 April 2015 J. Clin. Microbiol. doi:10.1128/JCM.00339-15 Copyright © 2015, American Society for Microbiology. All Rights Reserved.
1
First human case of meningitis and sepsis in a child caused by Actinobacillus suis/equuli
2
3
Carlotta Montagnania, Patrizia Pecileb, Maria Moriondoa, Patrizia Petriccic, Sabrina Becciania, Elena
4
Chiappinia, Giuseppe Indolfid, Gian Maria Rossolinib,e, f, Chiara Azzaria, Maurizio de Martinoa,
5
Luisa Gallia#
6
Department of Health Sciences, University of Florence and Anna Meyer Children's University
7
Hospital, Florence, Italya; Clinical Microbiology and Virology Unit, Department of Laboratory
8
Medicine, Careggi University Hospital, Florence, Italyb; Laboratory Medicine Unit, ASL6, Livorno,
9
Italyc; Anna Meyer Children's University Hospitald; Department of Experimental and Clinical
10
Medicine, University of Florencee; Department of Medical Biotechnologies, University of Sienaf
11
12
Running Head: First human case of A. suis/equuli invasive infection.
13
14
#Address correspondence to Professor Luisa Galli, MD, e-mail address: [email protected].
1
15
ABSTRACT
16
We report the first human case of meningitis and sepsis in a child caused by Actinobacillus suis or
17
equuli, common opportunistic pathogens of swine or horses. Identification was performed by
18
MALDI-TOF mass spectrometry and real-time PCR assay. Previous visit to a farm was suspected
19
as a possible source of infection.
2
20
CASE REPORT
21
A previously healthy 13-year-old boy who had been suffering since 24 hours from otalgia,
22
developed fever, headache, photophobia, drowsiness and neck stiffness, and was conducted to the
23
Emergency Department of a local hospital. No cranial traumas were reported. No other meningeal
24
signs, neurological deficits or papilledema were present at physical examination. Otoscopic findings
25
were normal. No petechiae were revealed. Laboratory tests showed neutrophilic leukocytosis (white
26
blood cells [WBC]: 17,700/μL; neutrophils [N]: 85%) and C-reactive protein (CRP):1.0 mg/dL. For
27
worsening drowsiness, an non-contrast-enhanced magnetic resonance of the head and neck was
28
performed that was normal. A lumbar puncture yielded an opalescent cerebrospinal fluid (CSF)
29
under normal pressure, with 650 WBC/μL, mainly polymorphonuclears, increased total protein (254
30
mg/dL) and normal glucose levels (49 mg/dL). Serum glucose level was 113 mg/dl. The patient was
31
given intravenous ceftriaxone (50 mg/kg) as empiric treatment and was transferred to our tertiary
32
referral Children's University Hospital. Laboratory tests were repeated and showed increased
33
neutrophilic leukocytosis (WBC: 38,560/μL, N: 95%) and increased CRP (12.7 mg/dL). A CSF
34
Gram stain, performed at the local hospital, revealed a Gram-negative bacillus. Cefriaxone was then
35
switched to meropenem 40 mg/kg/dose three times per day, and dexamethasone was added at 0.1
36
mg/kg/dose four times per day.
37
Blood cultures, carried out using the Bactec system with aerobic vials (Becton Dickinson, Milan,
38
Italy) were positive after 24 hours for a Gram-negative bacillus. The CSF culture yielded growth of
39
several colonies of a Gram-negative bacillus on Columbia blood agar and Chocolate agar
40
supplemented with PolyViteX (bioMérieux, Florence, Italy) after incubation for 24 h at 37°C under
41
aerobic conditions, while no growth was present on MacConkey agar. The bacterial isolates were
42
identified by the Vitek2 system (bioMérieux) as Actinobacillus ureae (reported as excellent
3
43
identification). However, the same isolates were identified as Actinobacillus suis/equuli by
44
MALDI-TOF mass spectrometry (Vitek MS, bioMérieux) with an identification score of 99.9%.
45
The real-time PCR tests for Neisseria meningitidis, Streptococcus pneumoniae, Haemophilus
46
influenzae type B and C, H. influenzae non-capsulated, Escherichia coli, Klebsiella pneumonie,
47
Streptocuccus agalactiae, Listeria monocitogenes performed on DNA extract from CSF using
48
Eusepscreen kit (Eurospital, Italy) resulted negative.
49
The 16S rDNA sequence, 997 bp long, confirmed the presence of Actinobacillus spp.(1). To
50
discriminate between A. ureae and A. suis/equuli a real-time PCR test was performed. The real-
51
time PCR assay amplifies the hypervariable region of 23S rRNA gene specific for A. suis (2) and,
52
as recently reported, for A. equuli. The 23S rRNA gene sequences were obtained from the NCBI
53
GenBank Database (EU333989.1, CP007715.1). The real-time PCR assay was performed in 25 μL
54
reaction volumes containing 2× TaqMan Universal Master Mix (Applied Biosystems); primers were
55
used at a concentration of 300 nM; the FAM-labelled probe was used at a concentration of 25 nM.
56
Six μL of DNA were used for each reaction. All reactions were performed in triplicate. DNA was
57
amplified in an ABI 7500 sequence detection system (Applied Biosystems) using the following
58
cycling parameters: 95 °C for 10 min followed by 45 cycles of a two-stage temperature profile of 95
59
°C for 15 s and 60 °C for 1 min. Primer and probe (analyzed using the Primer Express 2.0 program)
60
are shown in table 1.
61
Antimicrobial susceptibility was determined by microdilution broth method and results interpreted
62
by EUCAST PK/PD (non-species related) breakpoints (http://www.eucast.org, Table v5.0). Data
63
are shown in table 2.
64
We carried out a new careful history which revealed that the patient had visited a farm three days
65
before the onset of symptoms, although close contacts with horses or swine were denied. 4
66
Two days later, the patient’s condition improved, with fever resolution. Headache, neck stiffness
67
and drowsiness subsided in four days. Dexamethasone was reduced and discontinued in one week.
68
The child was discharged after 12 days of treatment and a 6-month follow-up examination did not
69
show any signs and symptoms of disease. A complete audiologic evaluation was performed that
70
was normal. T and B cell subsets, immunoglobulin levels and neutrophil function were evaluated
71
and were completely normal.
72
------------------------------------------------------------------------------------------------------------
73
We report a case of sepsis and meningitis due to A. suis/equuli occurred in a previously healthy
74
13 year-old boy. To the best of our knowledge, this is the first description of A. suis/equuli invasive
75
infection in children.
76
Actinobacillus is a Gram-negative coccobacillus, member of the Pasteurellaceae family. Humans
77
can be rarely colonized or infected by Actinobacillus hominis or A. ureae (3, 4). A. hominis and A.
78
ureae have been reported as uncommon commensal of the upper respiratory tract and can rarely
79
cause severe infections in humans (3, 4). Twenty-seven cases of infection, including 14 cases of
80
meningitis due to A. ureae in humans have been reported in the literature (5). A. ureae meningitis
81
mainly occurred in immunocompromised patients or after skull injury (4).
82
A. suis is an opportunistic pathogen of swine. A. suis is an early colonizer of upper respiratory
83
airways of swine and can also cause a wide range of invasive infections, including arthritis,
84
pleuropneumonia, septicemia and meningitis in pigs of all ages (6, 7, 8). A. equuli is a commensal
85
of horses and is a common cause of septicemia in foals (9). A. suis and A. equuli have been reported
86
to cause wound infection in humans after pig or horse bites (10, 11).A case of A. equuli septicemia
87
has been described in a 53-year-old butcher after a cut (12). However, A. suis or A. equuli have
88
never been reported to cause meningitis and sepsis in children. The most likely source of A. 5
89
suis/equuli could have been the farm visited by the child three days before the onset of symptoms,
90
although physical contacts with swine or horses was denied.
91
The patient presented sepsis and the hallmark symptoms of meningitis: fever, headache,
92
photophobia, neck stiffness and severe drowsiness. No alteration of coagulation were observed. No
93
characteristic signs and symptoms that allow to suspect a A. suis/equuli infection were identified.
94
Empirical treatment with ceftriaxone was started as soon as meningitis was suspected, following an
95
international guideline (13). However, when the Gram stain revealed a Gram-negative bacillus, an
96
Escherichia coli infection was suspected. Therefore, as there is a high prevalence of extended
97
spectrum β-lactamase producing bacteria is reported in our country (14), a switch to meropenem
98
was decided. Nevertheless, A. suis/equuli antimicrobial susceptibility showed low MICs for all
99
antibiotics tested, including cephalosporins and carbapenems.
100
Clinical response was optimal and the patient recovered without any sequelae.
101
When A. suis/equuli was identified, an immunological defect was suspected. A careful
102
immunological evaluation, including T and B-cell subsets, immunoglobulin levels and
103
immunoglobulin G subclasses, and neutrophil function was performed and the immunological
104
profile was completely normal. Moreover, the patient had experienced no other significant
105
condition, suggestive of immunological defects, in the past.
106
As identification with conventional biochemical profiling by Vitek2 yielded an identification of A.
107
ureae, while identification of A. suis/equuli could only be suspected using MALDI-TOF and
108
confirmed by molecular identification, we suggest that at least some of the previously reported
109
invasive infections by A. ureae could have been caused by A. suis/equuli.
110
In conclusion, we report the first case of A. suis/equuli sepsis and meningitis in children. We did not
111
identify any clinical characteristics that allow to suspect A. suis/equuli infection. Misdiagnosis of 6
112
this pathogen could be possible by the Vitek2 system. MALDI-TOF mass spectrometry and
113
molecular biology techniques may provide the proper diagnosis of this uncommon pathogen.
114
115
ACKNOWLEDGMENTS
116
Authors do not have conflicts of interest to report.
7
117
118
REFERENCES 1.
Lu JJ, Perng CL, Lee SY, Wan CC. 2000. Use of PCR with universal primers and
119
restriction endonuclease digestions for detection and identification of common bacterial
120
pathogens in cerebrospinal fluid. J. Clin. Microbiol. 38:2076-80.
121
2.
Kariyawasam S, Strait E, Jordan D, Kroll J. 2011. Development of a real-time
122
polymerase chain reaction assay for detection of Actinobacillus suis in porcine lung. J. Vet.
123
Diagn. Invest. 23:885-889.
124
3.
Friis-Møller A, Christensen JJ, Fussing V, Hesselbjerg A, Christiansen J, Bruun B.
125
2001. Clinical significance and taxonomy of Actinobacillus hominis. J. Clin. Microbiol.
126
39:930-935.
127
4.
of the literature. J. Emerg. Med. 13:623-627.
128 129
Kingsland RC, Guss DA. 1995. Actinobacillus ureae meningitis: case report and review
5.
de Castro N, Pavie J, Lagrange-Xélot M, Bouvry D, Delisle F, Parrot A, Molina JM.
130
2007. Severe Actinobacillus ureae meningitis in an immunocompromised patient: report of
131
one case and review of the literature. Scand. J. Infect. Dis. 39:1076-1079.
132
6.
MacInnes JI, Mackinnon J, Bujold AR, Ziebell K, Kropinski AM, Nash JH. 2012.
133
Complete genome sequence of Actinobacillus suis H91-0380, a virulent serotype O2 strain.
134
J. Bacteriol. 194:6686-6687.
135
7.
MacInnes JI, Desrosiers R. 1999. Agents of the "suis-ide diseases" of swine:
136
Actinobacillus suis, Haemophilus parasuis, and Streptococcus suis. Can. J. Vet. Res. 63:83-
137
89.
138
8.
colonization-deficient mutants of Actinobacillus suis. Vet. Microbiol. 140:122-130.
139 140 141
Ojha S, Lacouture S, Gottschalk M, MacInnes JI. 2010. Characterization of
9.
Thompson AB, Postey RC, Snider T, Pasma T. 2010. Actinobacillus equuli as a primary pathogen in breeding sows and piglets. Can. Vet. J. 51:1223-1225. 8
142
10.
bite. Lancet. 348:888.
143 144
Escande F, Bailly A, Bone S, Lemozy J. 1996. Actinobacillus suis infection after a pig
11.
Peel MM, Hornidge KA, Luppino M, Stacpoole AM, Weaver RE. 1991.
145
Actinobacillus spp. and related bacteria in infected wounds of humans bitten by horses and
146
sheep. J. Clin. Microbiol. 29:2535-2538.
147
12.
septicemia: an unusual zoonotic infection. J. Clin. Microbiol. 36:2789-2790.
148 149
Ashhurst-Smith C, Norton R, Thoreau W, Peel MM. 1998. Actinobacillus equuli
13.
National Institute for Health and Clinical Excellence. 2010. Bacterial meningitis and
150
meningococcal septicaemia. Management of bacterial meningitis and meningococcal
151
septicaemia in children and young people younger than 16 years in primary and secondary
152
care. NICE clinical guideline 102. URL:guidance.nice.org.uk/cg102.
153
14.
European Centre for Disease Prevention and Control. 2013. Antimicrobial resistance
154
surveillance in Europe 2012. Annual Report of the European Antimicrobial Resistance
155
Surveillance Network (EARS-Net). ECDC. doi:10.2900/93403.
156 157 158
9
159
TABLE 1: Primer and probe sequences Sequence (5′ to 3′) Gene* Forward primer 23S-rRNA
299
-CCG TAT TCG AAG TGT CTA TTG TGG TA-324
* GenBank accession no. EU333989.1
Reverse primer 385
-GAT GGT CCC CCC ATC TTC A-367
Probe FAM 332-AAC GAC AAG TAG GGC GGG ACA CGA-355 TAMRA FAM
6-carboxyfluorescein
TAMRA
tetramethylrhodamine
10
160
TABLE 2: Antimicrobial susceptibility of the Actinobacillus isolate Antibiotic
MIC (μg/ml)
Susceptibility
EUCAST 5.0a amoxicillin/clavulanic acid (2:1 ratio)
CLSIb
1
First human case of meningitis and sepsis in a child caused by Actinobacillus suis/equuli
2
3
Carlotta Montagnania, Patrizia Pecileb, Maria Moriondoa, Patrizia Petriccic, Sabrina Becciania, Elena
4
Chiappinia, Giuseppe Indolfid, Gian Maria Rossolinib,e, f, Chiara Azzaria, Maurizio de Martinoa,
5
Luisa Gallia#
6
Department of Health Sciences, University of Florence and Anna Meyer Children's University
7
Hospital, Florence, Italya; Clinical Microbiology and Virology Unit, Department of Laboratory
8
Medicine, Careggi University Hospital, Florence, Italyb; Laboratory Medicine Unit, ASL6, Livorno,
9
Italyc; Anna Meyer Children's University Hospitald; Department of Experimental and Clinical
10
Medicine, University of Florencee; Department of Medical Biotechnologies, University of Sienaf
11
12
Running Head: First human case of A. suis/equuli invasive infection.
13
14
#Address correspondence to Professor Luisa Galli, MD, e-mail address: [email protected].
1
15
ABSTRACT
16
We report the first human case of meningitis and sepsis in a child caused by Actinobacillus suis or
17
equuli, common opportunistic pathogens of swine or horses. Identification was performed by
18
MALDI-TOF mass spectrometry and real-time PCR assay. Previous visit to a farm was suspected
19
as a possible source of infection.
2
20
CASE REPORT
21
A previously healthy 13-year-old boy who had been suffering since 24 hours from otalgia,
22
developed fever, headache, photophobia, drowsiness and neck stiffness, and was conducted to the
23
Emergency Department of a local hospital. No cranial traumas were reported. No other meningeal
24
signs, neurological deficits or papilledema were present at physical examination. Otoscopic findings
25
were normal. No petechiae were revealed. Laboratory tests showed neutrophilic leukocytosis (white
26
blood cells [WBC]: 17,700/μL; neutrophils [N]: 85%) and C-reactive protein (CRP):1.0 mg/dL. For
27
worsening drowsiness, an non-contrast-enhanced magnetic resonance of the head and neck was
28
performed that was normal. A lumbar puncture yielded an opalescent cerebrospinal fluid (CSF)
29
under normal pressure, with 650 WBC/μL, mainly polymorphonuclears, increased total protein (254
30
mg/dL) and normal glucose levels (49 mg/dL). Serum glucose level was 113 mg/dl. The patient was
31
given intravenous ceftriaxone (50 mg/kg) as empiric treatment and was transferred to our tertiary
32
referral Children's University Hospital. Laboratory tests were repeated and showed increased
33
neutrophilic leukocytosis (WBC: 38,560/μL, N: 95%) and increased CRP (12.7 mg/dL). A CSF
34
Gram stain, performed at the local hospital, revealed a Gram-negative bacillus. Cefriaxone was then
35
switched to meropenem 40 mg/kg/dose three times per day, and dexamethasone was added at 0.1
36
mg/kg/dose four times per day.
37
Blood cultures, carried out using the Bactec system with aerobic vials (Becton Dickinson, Milan,
38
Italy) were positive after 24 hours for a Gram-negative bacillus. The CSF culture yielded growth of
39
several colonies of a Gram-negative bacillus on Columbia blood agar and Chocolate agar
40
supplemented with PolyViteX (bioMérieux, Florence, Italy) after incubation for 24 h at 37°C under
41
aerobic conditions, while no growth was present on MacConkey agar. The bacterial isolates were
42
identified by the Vitek2 system (bioMérieux) as Actinobacillus ureae (reported as excellent
3
43
identification). However, the same isolates were identified as Actinobacillus suis/equuli by
44
MALDI-TOF mass spectrometry (Vitek MS, bioMérieux) with an identification score of 99.9%.
45
The real-time PCR tests for Neisseria meningitidis, Streptococcus pneumoniae, Haemophilus
46
influenzae type B and C, H. influenzae non-capsulated, Escherichia coli, Klebsiella pneumonie,
47
Streptocuccus agalactiae, Listeria monocitogenes performed on DNA extract from CSF using
48
Eusepscreen kit (Eurospital, Italy) resulted negative.
49
The 16S rDNA sequence, 997 bp long, confirmed the presence of Actinobacillus spp.(1). To
50
discriminate between A. ureae and A. suis/equuli a real-time PCR test was performed. The real-
51
time PCR assay amplifies the hypervariable region of 23S rRNA gene specific for A. suis (2) and,
52
as recently reported, for A. equuli. The 23S rRNA gene sequences were obtained from the NCBI
53
GenBank Database (EU333989.1, CP007715.1). The real-time PCR assay was performed in 25 μL
54
reaction volumes containing 2× TaqMan Universal Master Mix (Applied Biosystems); primers were
55
used at a concentration of 300 nM; the FAM-labelled probe was used at a concentration of 25 nM.
56
Six μL of DNA were used for each reaction. All reactions were performed in triplicate. DNA was
57
amplified in an ABI 7500 sequence detection system (Applied Biosystems) using the following
58
cycling parameters: 95 °C for 10 min followed by 45 cycles of a two-stage temperature profile of 95
59
°C for 15 s and 60 °C for 1 min. Primer and probe (analyzed using the Primer Express 2.0 program)
60
are shown in table 1.
61
Antimicrobial susceptibility was determined by microdilution broth method and results interpreted
62
by EUCAST PK/PD (non-species related) breakpoints (http://www.eucast.org, Table v5.0). Data
63
are shown in table 2.
64
We carried out a new careful history which revealed that the patient had visited a farm three days
65
before the onset of symptoms, although close contacts with horses or swine were denied. 4
66
Two days later, the patient’s condition improved, with fever resolution. Headache, neck stiffness
67
and drowsiness subsided in four days. Dexamethasone was reduced and discontinued in one week.
68
The child was discharged after 12 days of treatment and a 6-month follow-up examination did not
69
show any signs and symptoms of disease. A complete audiologic evaluation was performed that
70
was normal. T and B cell subsets, immunoglobulin levels and neutrophil function were evaluated
71
and were completely normal.
72
------------------------------------------------------------------------------------------------------------
73
We report a case of sepsis and meningitis due to A. suis/equuli occurred in a previously healthy
74
13 year-old boy. To the best of our knowledge, this is the first description of A. suis/equuli invasive
75
infection in children.
76
Actinobacillus is a Gram-negative coccobacillus, member of the Pasteurellaceae family. Humans
77
can be rarely colonized or infected by Actinobacillus hominis or A. ureae (3, 4). A. hominis and A.
78
ureae have been reported as uncommon commensal of the upper respiratory tract and can rarely
79
cause severe infections in humans (3, 4). Twenty-seven cases of infection, including 14 cases of
80
meningitis due to A. ureae in humans have been reported in the literature (5). A. ureae meningitis
81
mainly occurred in immunocompromised patients or after skull injury (4).
82
A. suis is an opportunistic pathogen of swine. A. suis is an early colonizer of upper respiratory
83
airways of swine and can also cause a wide range of invasive infections, including arthritis,
84
pleuropneumonia, septicemia and meningitis in pigs of all ages (6, 7, 8). A. equuli is a commensal
85
of horses and is a common cause of septicemia in foals (9). A. suis and A. equuli have been reported
86
to cause wound infection in humans after pig or horse bites (10, 11).A case of A. equuli septicemia
87
has been described in a 53-year-old butcher after a cut (12). However, A. suis or A. equuli have
88
never been reported to cause meningitis and sepsis in children. The most likely source of A. 5
89
suis/equuli could have been the farm visited by the child three days before the onset of symptoms,
90
although physical contacts with swine or horses was denied.
91
The patient presented sepsis and the hallmark symptoms of meningitis: fever, headache,
92
photophobia, neck stiffness and severe drowsiness. No alteration of coagulation were observed. No
93
characteristic signs and symptoms that allow to suspect a A. suis/equuli infection were identified.
94
Empirical treatment with ceftriaxone was started as soon as meningitis was suspected, following an
95
international guideline (13). However, when the Gram stain revealed a Gram-negative bacillus, an
96
Escherichia coli infection was suspected. Therefore, as there is a high prevalence of extended
97
spectrum β-lactamase producing bacteria is reported in our country (14), a switch to meropenem
98
was decided. Nevertheless, A. suis/equuli antimicrobial susceptibility showed low MICs for all
99
antibiotics tested, including cephalosporins and carbapenems.
100
Clinical response was optimal and the patient recovered without any sequelae.
101
When A. suis/equuli was identified, an immunological defect was suspected. A careful
102
immunological evaluation, including T and B-cell subsets, immunoglobulin levels and
103
immunoglobulin G subclasses, and neutrophil function was performed and the immunological
104
profile was completely normal. Moreover, the patient had experienced no other significant
105
condition, suggestive of immunological defects, in the past.
106
As identification with conventional biochemical profiling by Vitek2 yielded an identification of A.
107
ureae, while identification of A. suis/equuli could only be suspected using MALDI-TOF and
108
confirmed by molecular identification, we suggest that at least some of the previously reported
109
invasive infections by A. ureae could have been caused by A. suis/equuli.
110
In conclusion, we report the first case of A. suis/equuli sepsis and meningitis in children. We did not
111
identify any clinical characteristics that allow to suspect A. suis/equuli infection. Misdiagnosis of 6
112
this pathogen could be possible by the Vitek2 system. MALDI-TOF mass spectrometry and
113
molecular biology techniques may provide the proper diagnosis of this uncommon pathogen.
114
115
ACKNOWLEDGMENTS
116
Authors do not have conflicts of interest to report.
7
117
118
REFERENCES 1.
Lu JJ, Perng CL, Lee SY, Wan CC. 2000. Use of PCR with universal primers and
119
restriction endonuclease digestions for detection and identification of common bacterial
120
pathogens in cerebrospinal fluid. J. Clin. Microbiol. 38:2076-80.
121
2.
Kariyawasam S, Strait E, Jordan D, Kroll J. 2011. Development of a real-time
122
polymerase chain reaction assay for detection of Actinobacillus suis in porcine lung. J. Vet.
123
Diagn. Invest. 23:885-889.
124
3.
Friis-Møller A, Christensen JJ, Fussing V, Hesselbjerg A, Christiansen J, Bruun B.
125
2001. Clinical significance and taxonomy of Actinobacillus hominis. J. Clin. Microbiol.
126
39:930-935.
127
4.
of the literature. J. Emerg. Med. 13:623-627.
128 129
Kingsland RC, Guss DA. 1995. Actinobacillus ureae meningitis: case report and review
5.
de Castro N, Pavie J, Lagrange-Xélot M, Bouvry D, Delisle F, Parrot A, Molina JM.
130
2007. Severe Actinobacillus ureae meningitis in an immunocompromised patient: report of
131
one case and review of the literature. Scand. J. Infect. Dis. 39:1076-1079.
132
6.
MacInnes JI, Mackinnon J, Bujold AR, Ziebell K, Kropinski AM, Nash JH. 2012.
133
Complete genome sequence of Actinobacillus suis H91-0380, a virulent serotype O2 strain.
134
J. Bacteriol. 194:6686-6687.
135
7.
MacInnes JI, Desrosiers R. 1999. Agents of the "suis-ide diseases" of swine:
136
Actinobacillus suis, Haemophilus parasuis, and Streptococcus suis. Can. J. Vet. Res. 63:83-
137
89.
138
8.
colonization-deficient mutants of Actinobacillus suis. Vet. Microbiol. 140:122-130.
139 140 141
Ojha S, Lacouture S, Gottschalk M, MacInnes JI. 2010. Characterization of
9.
Thompson AB, Postey RC, Snider T, Pasma T. 2010. Actinobacillus equuli as a primary pathogen in breeding sows and piglets. Can. Vet. J. 51:1223-1225. 8
142
10.
bite. Lancet. 348:888.
143 144
Escande F, Bailly A, Bone S, Lemozy J. 1996. Actinobacillus suis infection after a pig
11.
Peel MM, Hornidge KA, Luppino M, Stacpoole AM, Weaver RE. 1991.
145
Actinobacillus spp. and related bacteria in infected wounds of humans bitten by horses and
146
sheep. J. Clin. Microbiol. 29:2535-2538.
147
12.
septicemia: an unusual zoonotic infection. J. Clin. Microbiol. 36:2789-2790.
148 149
Ashhurst-Smith C, Norton R, Thoreau W, Peel MM. 1998. Actinobacillus equuli
13.
National Institute for Health and Clinical Excellence. 2010. Bacterial meningitis and
150
meningococcal septicaemia. Management of bacterial meningitis and meningococcal
151
septicaemia in children and young people younger than 16 years in primary and secondary
152
care. NICE clinical guideline 102. URL:guidance.nice.org.uk/cg102.
153
14.
European Centre for Disease Prevention and Control. 2013. Antimicrobial resistance
154
surveillance in Europe 2012. Annual Report of the European Antimicrobial Resistance
155
Surveillance Network (EARS-Net). ECDC. doi:10.2900/93403.
156 157 158
9
159
TABLE 1: Primer and probe sequences Sequence (5′ to 3′) Gene* Forward primer 23S-rRNA
299
-CCG TAT TCG AAG TGT CTA TTG TGG TA-324
* GenBank accession no. EU333989.1
Reverse primer 385
-GAT GGT CCC CCC ATC TTC A-367
Probe FAM 332-AAC GAC AAG TAG GGC GGG ACA CGA-355 TAMRA FAM
6-carboxyfluorescein
TAMRA
tetramethylrhodamine
10
160
TABLE 2: Antimicrobial susceptibility of the Actinobacillus isolate Antibiotic
MIC (μg/ml)
Susceptibility
EUCAST 5.0a amoxicillin/clavulanic acid (2:1 ratio)
CLSIb
