JCM Accepts, published online ahead of print on 16 July 2014 J. Clin. Microbiol. doi:10.1128/JCM.00726-14 Copyright © 2014, American Society for Microbiology. All Rights Reserved.
1
Title:
2
First American Isolate of KPC-2 Producing Klebsiella pneumonaie Sequence Type 23
3 4
Authors:
5
Daniela Cejasa, Liliana Fernández Canigiab, Giovanna Rincón Cruza, c, AlanX.Elenaa, Ivana
6
Maldonadob, GabrielO.Gutkinda, Marcela A. Radice#,a.
7 8
Affiliations:
9
Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Argentinaa; Hospital
10
Alemán, Ciudad Autónoma de Buenos Aires, Argentinab; Escuela de Bacteriología,
11
Universidad Industrial de Santander, Bucaramanga, Colombiac.
12 13
Running title: KPC-2 producing Klebsiella pneumonaie ST23
14 15
Corresponding author: Radice Marcela
16
Mail: [email protected]
17 18
1
19
Abstract:
20
KPC-2 producing Klebsiella pneumoniae mainly correspond to the CC258, however we
21
describe KPC-2 producing K. pneumoniae belonging to the invasive ST23. KPC-2 has been
22
scarce reported in ST23 and this constitutes its first isolation in America. Acquisition of
23
resistant markers in virulent clones could mark an evolutionary step towards their settling
24
as major nosocomial pathogen.
25 26
Case report:
27
An elderly, 85-year-old, man was admitted at the Intensive Care Unit of a hospital at
28
Buenos Aires, in March 19th, 2013. He presented poor general condition, sensory
29
impairment, hypotension, poor peripheral perfusion, crackling rales and desaturation. He
30
has history of acute myeloid leukemia in 2012, currently undergoing chemotherapy with
31
methotrexate (20 mg/week) and prednisone (150 mg/day). Two days after his admission a
32
meticillin susceptible Staphylococcus aureus was isolated from blood culture and tracheal
33
aspirate (104 UFC/ml) and the patient was treated with cefazolin. A week later, the patient
34
developed a catheter associated bacteremia due to meticillin resistant S. epidermidis and he
35
received linezolid. He presented intercurrent hypovolemic shock and hypotension requiring
36
transfusion of 2 units of red blood cells. Concurrently a hypermucoviscous Klebsiella
37
pneumoniae (K. penumoniae 3089) was recovered from a second tracheal aspirate culture
38
(105 UFC/ml). The general condition of the patient worsened and finally died on April
39
19th.
40
Antimicrobial susceptibility tests were conducted on K. pneumoniae 3089 according to
41
CLSI (1). The isolate was resistant to all beta-lactams, including carbapenems, but
42
remained
susceptible
to
aminoglycosides,
fluoroquinolones,
trimethoprime 2
43
sulfamethoxazole, doxycycline, fosfomycin, colistin and tigecyclin. A positive synergy test
44
between imipenem (30 μg) and phenyl boronic acid (300 μg) containing disks indicated the
45
possible presence of KPC beta-lactamases. The presence of blaKPC was confirmed by PCR
46
amplification and its genetic context was investigated by PCR mapping and sequencing,
47
using plasmid DNA as template (figure 1) (2). As expected, blaKPC-2 was located in Tn4401
48
as previously reported by Naas et al (3). Replicon typing, determined according to Carattoli
49
et al (4), indicated that blaKPC-2 containing plasmid corresponded to FIA Inc group, which
50
had been previously reported in E. coli, in Argentina, by Gomez et al (5). Conjugation
51
assays, using both Escherichia coli HB101 and Escherichia coli CAG 12177 as receptor
52
strains, did not yield transconjugants, according to the previously mentioned study (5).
53
KPC-producing K. pneumoniae are, nowadays, endemic in different countries. The
54
successful dissemination of K. pneumoniae belonging to the clonal complex 258 was a
55
critical factor resulting in their pandemic expansion (6). In our country, a substantial
56
increase of KPC-2 producing K. pneumoniae was observed in 2010, due to the huge
57
dissemination of the hyperendemic ST 258 clone, which displayed a multidrug resistant
58
phenotype (5, 7, 8). A multilocus sequence typing scheme (MLST) was conducted on K.
59
pneumoniae 3089 (9). Unexpectedly, it displayed the following allelic profile: gapA 2, infB
60
1, mdh 1, pgi 1, phoE 9, rpoB 4, tonB12, corresponding to ST23.
61
K. pneumoniae belonging to ST23 correspond to a hypermucoviscous phenotype.
62
Hypermucoviscous strains are associated to a highly invasive syndrome characterized by
63
bacteremia, liver abscesses, metastatic infections, and even endophthalmitis, supurative
64
meningitis and brain abscess (10, 11). The invasive nature of K. pneumoniae ST23 seems to
65
correlate with the hypermucoviscosity that protects from phagocytosis and serum killing by
66
complement. The plasmid-mediated regulator of mucoid phenotype A (rmpA) and the 3
67
mucoviscosity-associated gene A (magA) have been associated to this virulent phenotype
68
(12-14). The last, renamed as wzyKpK1, is a chromosomal gene that is required for
69
exopolysaccharide biosynthesis and is restricted to K. pneumoniae capsule serotype K1,
70
considered to be the most virulent strains of K. pneumoniae (13). Most of the isolates from
71
patients with K. pneumoniae liver abscess syndrome (KLAS) belong to the K1 serotype and
72
correspond to ST23 (14, 15). Although KLAS are endemic in Taiwan, they have been
73
reported with increasing frequency in other countries in Southeast Asia. They constitute an
74
emerging infectious disease in the United States and Europe, moreover they were recently
75
reported in Argentina (11, 13, 16). Hypermucoviscous K. pneumoniae isolates, including
76
ST23 clinical strains, have been found to be susceptible to most antibiotics, including third-
77
and fourth-generation cephalosporins, monobactams, carbapenems and ciprofloxacin (17).
78
As K. pneumoniae 3089 exhibited an extreme colony stickiness and rendered a positive
79
string test (18) (figure 2), the presence of magA and rpmA virulence genes was investigated
80
using the following primers (5´-3´): wzy-F: CGCCGCAAATACGAGAAGTG, wzy-R:
81
GCAATCGAAGTGAAGAGTGC and rmpA-F: ACTGGGCTACCTCTGCTTCA, rmpA-
82
R: CTTGCATGAGCCATCTTTCA. Both hypermucoviscocity associated genes were
83
detected in the studied isolate.
84
Although K. pneumoniae ST23 isolates usually display a susceptible profile, here we
85
detected the presenceof KPC-2 in an isolate belonging to this invasive sequencetype. The
86
presence of KPC carbapenemases in K. pneumoniae ST23 has been only previously
87
reported in isolates from China and Poland, in 2010 and 2011, respectively, displaying the
88
same susceptibility profile as K. pneumoniae 3089 (17, 19, 20). However, no single
89
mention to the virulence factors or hypermucoviscosity phenotype was included in those
90
studies. 4
91
In the last few months three more hypermucoviscous K. pneumoniae ST23 isolates have
92
been referred to our laboratory, displaying antimicrobial susceptible phenotypes except for
93
ampicillin. Considering the virulence factors associated to this phenotype and its highly
94
invasive feature a prompt identification and accurate treatment should be mandatory. These
95
strains can be readily detected by the string test, MLST and molecular characterization of
96
the hypermucoviscous phenotype associated genes. Antibiotics commonly used in K.
97
pneumoniae infections have been useful for the therapeutic treatment of ST23 clinical
98
isolates, however the acquisition of resistance genes in this invasive strains could hinder
99
their eradication, probably favoring the development of metastatic infections.
100
A rising number of cases of K. pneumoniae ST23 from geographic regions other than
101
Southeast Asia indicate that this is a globally emerging pathogen. According to Brisse et
102
al., K. pneumoniae ST23 constitutes an emerging highly virulent and metabolically
103
versatile clone (14), so the acquisition of an important mechanism of antibiotic resistance
104
such as KPC-2 could mark an evolutionary step towards the settling of K. pneumoniae
105
ST23 as a major cause of nosocomial infections.
106 107
Acknowledgments
108
This work was partially supported by Grants from UBACyT and ANPCyT to M. Radice
109
and G. Gutkind. G. Gutkind and M. Radice are members of Carrera del Investigador
110
Científico (CONICET). D. Cejas was recipientof a doctoral fellowship from CONICET and
111
is now recipient of a post doctoral fellowship from Fundación Bunge y Born.
112 113 114 5
115
References
116
1.
117
antimicrobial susceptibility testing; twenty-first informational supplement M100-S23.
118
2013;33 (1).
119
2.
120
Brooks S, Cebular S, Quale J. 2004. Emergence of carbapenem-resistant Klebsiella
121
species possessing the class A carbapenem-hydrolyzing KPC-2 and inhibitor-resistant
122
TEM-30 beta-lactamases in New York City. Clin Infect Dis. 39:55-60.
123
3.
124
Genetic structures at the origin of acquisition of the beta-lactamase blaKPC gene.
125
Antimicrob Agents Chemother. 52:1257-1263.
126
4.
127
Identification of plasmids by PCR-based replicon typing. J Microbiol Methods. 63:219-
128
228.
129
5.
130
Lastovetska O, Albornoz E, Galas M, Melano RG, Corso A, Petroni A. 2011. Clonal
131
dissemination of Klebsiella pneumoniae ST258 harbouring KPC-2 in Argentina. Clin
132
Microbiol Infect. 17:1520-1524.
133
6.
134
carbapenemase-producing bacteria. Lancet Infect Dis. 9:228-236.
135
7.
136
Vay C, Maldonado I, Famiglietti A, Giovanakis M, Magarinos F, Berardinelli E,
137
Neira L, Mollerach M, Gutkind G, Radice M. 2012. Hyperendemic clone of KPC
Clinical and Laboratory Standards Institute. Performance standards for
Bradford PA, Bratu S, Urban C, Visalli M, Mariano N, Landman D, Rahal, JJ,
Naas T, Cuzon G, Villegas MV, Lartigue MF, Quinn JP, Nordmann P. 2008.
Carattoli A, Bertini A, Villa L, Falbo V, Hopkins KL, Threlfall EJ. 2005.
Gomez SA, Pasteran FG, Faccone D, Tijet N, Rapoport M, Lucero C,
Nordmann P, Cuzon G, Naas T. 2009. The real threat of Klebsiella pneumoniae
Cejas D, Fernandez Canigia L, Nastro M, Rodriguez C, Tanco A, Rodriguez H,
6
138
producing Klebsiella pneumoniae ST 258 in Buenos Aires hospitals. Infect Genet Evol.
139
12:499-501
140
8.
141
Faccone D, Di Martino A, Galas M. 2008. Klebsiella pneumoniae Carbapenemase-2,
142
Buenos Aires, Argentina. Emerg Infect Dis. 14:1178-1180.
143
9.
144
sequence typing of Klebsiella pneumoniae nosocomial isolates. J Clin Microbiol. 43:4178-
145
4182.
146
10.
147
with septic endophthalmitis. Arch Intern Med. 146:1913-1916.
148
11.
149
endemic disease, Taiwan. Emerg Infect Dis. 14:1592-1600.
150
12.
151
of colanic acid synthesis in Escherichia coli by rmpA and rmpB, two virulence-plasmid
152
genes of Klebsiella pneumoniae. Mol Microbiol. 3:1349-1359.
153
13.
154
the putative virulence gene magA in a worldwide collection of 495 Klebsiella isolates:
155
magA is restricted to the gene cluster of Klebsiella pneumoniae capsule serotype K1. J Med
156
Microbiol. 54:1111-1113.
157
14.
158
Grimont P. 2009. Virulent clones of Klebsiella pneumoniae: identification and
159
evolutionary scenario based on genomic and phenotypic characterization. PLoS
160
One.4:e4982.
Pasteran FG, Otaegui L, Guerriero L, Radice G, Maggiora R, Rapoport M,
Diancourt L, Passet V, Verhoef J, Grimont PA, Brisse S. 2005. Multilocus
Liu YC, Cheng DL, Lin CL. 1986. Klebsiella pneumoniae liver abscess associated
Tsai FC, Huang YT, Chang LY, Wang JT. 2008. Pyogenic liver abscess as
Nassif X, Honore N, Vasselon T, Cole ST, Sansonetti PJ. 1989. Positive control
Struve C, Bojer M, Nielsen EM, Hansen DS, Krogfelt KA. 2005. Investigation of
Brisse S, Fevre C, Passet V, Issenhuth-Jeanjean S, Tournebize R, Diancourt L,
7
161
15.
Turton JF, Englender H, Gabriel SN, Turton SE, Kaufmann ME, Pitt TL.
162
2007. Genetically similar isolates of Klebsiella pneumoniae serotype K1 causing liver
163
abscesses in three continents. J Med Microbiol. 56:593-597.
164
16.
165
Appearance of Klebsiella pneumoniae liver abscess syndrome in Argentina: case report and
166
review of molecular mechanisms of pathogenesis. Open Microbiol J. 5:107-113.
167
17.
168
plasmid carrying blaKPC-2 in Enterobacteriaceae isolates from distinct cities in China.
169
Microb Drug Resist. 16:297-301.
170
18.
171
pneumoniae in North America. J Clin Microbiol. 43:991-992.
172
19.
173
D, Kania-Pudlo M, Mlynarczyk G, Zak-Pulawska Z, Hryniewicz W, Gniadkowski M.
174
2011. Molecular characteristics of KPC-producing Enterobacteriaceae at the early stage of
175
their dissemination in Poland, 2008-2009. Antimicrob Agents Chemother. 55:5493-5499.
176
20.
177
producing Klebsiella pneumoniae in China. J Antimicrob Chemother. 66:307-312.
Vila A, Cassata A, Pagella H, Amadio C, Yeh KM, Chang FY, Siu LK. 2011.
Qi Y, Wei Z, Li L, Ji S, Du X, Shen P, Yu Y. 2010. Detection of a common
Fang FC, Sandler N, Libby SJ. 2005. Liver abscess caused by magA+ Klebsiella
Baraniak A, Grabowska A, Izdebski R, Fiett J, Herda M, Bojarska K, Zabicka
Qi Y, Wei Z, Ji S, Du X, Shen P, Yu Y. 2011. ST11, the dominant clone of KPC-
178
8
179
Figure 1: Genetic context of blaKPC-2.
180
Legend figure 1: Primers (5’-3’) used to perform the PCR mapping of blaKPC-2. KPC-F:
181
ATGTCAC TGTATCGCCGTCT, KPC-R:TTTTCAGAGCCTTACTGCCC (2), 816U:
182
CACCTACACCACGACGAACC, 3098U: TGACCCTGAGCGGCGAAAGC and 4714:
183
GAAGATGCCAAGGTCAATGC (3) and IstB-RV:TTCCTGACCACTCCCGCCTTCC
184
(this study).
185 186
Figure 2: Hypermucoviscous phenotype of K. pneumoniae 3089
187
Legend figure 2: Hypermucoviscous phenotype is characterized by the formation of
188
elongated (≥5 mm) mucoviscous strings when a loop is passed through a colony (positive
189
string test).
9
1kb
t npA
I s t A 816U
I s t B
bl aKPC-2
3098U
t npA
KPCFW
I s t BRV
KPCRV
4714
1
Title:
2
First American Isolate of KPC-2 Producing Klebsiella pneumonaie Sequence Type 23
3 4
Authors:
5
Daniela Cejasa, Liliana Fernández Canigiab, Giovanna Rincón Cruza, c, AlanX.Elenaa, Ivana
6
Maldonadob, GabrielO.Gutkinda, Marcela A. Radice#,a.
7 8
Affiliations:
9
Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Argentinaa; Hospital
10
Alemán, Ciudad Autónoma de Buenos Aires, Argentinab; Escuela de Bacteriología,
11
Universidad Industrial de Santander, Bucaramanga, Colombiac.
12 13
Running title: KPC-2 producing Klebsiella pneumonaie ST23
14 15
Corresponding author: Radice Marcela
16
Mail: [email protected]
17 18
1
19
Abstract:
20
KPC-2 producing Klebsiella pneumoniae mainly correspond to the CC258, however we
21
describe KPC-2 producing K. pneumoniae belonging to the invasive ST23. KPC-2 has been
22
scarce reported in ST23 and this constitutes its first isolation in America. Acquisition of
23
resistant markers in virulent clones could mark an evolutionary step towards their settling
24
as major nosocomial pathogen.
25 26
Case report:
27
An elderly, 85-year-old, man was admitted at the Intensive Care Unit of a hospital at
28
Buenos Aires, in March 19th, 2013. He presented poor general condition, sensory
29
impairment, hypotension, poor peripheral perfusion, crackling rales and desaturation. He
30
has history of acute myeloid leukemia in 2012, currently undergoing chemotherapy with
31
methotrexate (20 mg/week) and prednisone (150 mg/day). Two days after his admission a
32
meticillin susceptible Staphylococcus aureus was isolated from blood culture and tracheal
33
aspirate (104 UFC/ml) and the patient was treated with cefazolin. A week later, the patient
34
developed a catheter associated bacteremia due to meticillin resistant S. epidermidis and he
35
received linezolid. He presented intercurrent hypovolemic shock and hypotension requiring
36
transfusion of 2 units of red blood cells. Concurrently a hypermucoviscous Klebsiella
37
pneumoniae (K. penumoniae 3089) was recovered from a second tracheal aspirate culture
38
(105 UFC/ml). The general condition of the patient worsened and finally died on April
39
19th.
40
Antimicrobial susceptibility tests were conducted on K. pneumoniae 3089 according to
41
CLSI (1). The isolate was resistant to all beta-lactams, including carbapenems, but
42
remained
susceptible
to
aminoglycosides,
fluoroquinolones,
trimethoprime 2
43
sulfamethoxazole, doxycycline, fosfomycin, colistin and tigecyclin. A positive synergy test
44
between imipenem (30 μg) and phenyl boronic acid (300 μg) containing disks indicated the
45
possible presence of KPC beta-lactamases. The presence of blaKPC was confirmed by PCR
46
amplification and its genetic context was investigated by PCR mapping and sequencing,
47
using plasmid DNA as template (figure 1) (2). As expected, blaKPC-2 was located in Tn4401
48
as previously reported by Naas et al (3). Replicon typing, determined according to Carattoli
49
et al (4), indicated that blaKPC-2 containing plasmid corresponded to FIA Inc group, which
50
had been previously reported in E. coli, in Argentina, by Gomez et al (5). Conjugation
51
assays, using both Escherichia coli HB101 and Escherichia coli CAG 12177 as receptor
52
strains, did not yield transconjugants, according to the previously mentioned study (5).
53
KPC-producing K. pneumoniae are, nowadays, endemic in different countries. The
54
successful dissemination of K. pneumoniae belonging to the clonal complex 258 was a
55
critical factor resulting in their pandemic expansion (6). In our country, a substantial
56
increase of KPC-2 producing K. pneumoniae was observed in 2010, due to the huge
57
dissemination of the hyperendemic ST 258 clone, which displayed a multidrug resistant
58
phenotype (5, 7, 8). A multilocus sequence typing scheme (MLST) was conducted on K.
59
pneumoniae 3089 (9). Unexpectedly, it displayed the following allelic profile: gapA 2, infB
60
1, mdh 1, pgi 1, phoE 9, rpoB 4, tonB12, corresponding to ST23.
61
K. pneumoniae belonging to ST23 correspond to a hypermucoviscous phenotype.
62
Hypermucoviscous strains are associated to a highly invasive syndrome characterized by
63
bacteremia, liver abscesses, metastatic infections, and even endophthalmitis, supurative
64
meningitis and brain abscess (10, 11). The invasive nature of K. pneumoniae ST23 seems to
65
correlate with the hypermucoviscosity that protects from phagocytosis and serum killing by
66
complement. The plasmid-mediated regulator of mucoid phenotype A (rmpA) and the 3
67
mucoviscosity-associated gene A (magA) have been associated to this virulent phenotype
68
(12-14). The last, renamed as wzyKpK1, is a chromosomal gene that is required for
69
exopolysaccharide biosynthesis and is restricted to K. pneumoniae capsule serotype K1,
70
considered to be the most virulent strains of K. pneumoniae (13). Most of the isolates from
71
patients with K. pneumoniae liver abscess syndrome (KLAS) belong to the K1 serotype and
72
correspond to ST23 (14, 15). Although KLAS are endemic in Taiwan, they have been
73
reported with increasing frequency in other countries in Southeast Asia. They constitute an
74
emerging infectious disease in the United States and Europe, moreover they were recently
75
reported in Argentina (11, 13, 16). Hypermucoviscous K. pneumoniae isolates, including
76
ST23 clinical strains, have been found to be susceptible to most antibiotics, including third-
77
and fourth-generation cephalosporins, monobactams, carbapenems and ciprofloxacin (17).
78
As K. pneumoniae 3089 exhibited an extreme colony stickiness and rendered a positive
79
string test (18) (figure 2), the presence of magA and rpmA virulence genes was investigated
80
using the following primers (5´-3´): wzy-F: CGCCGCAAATACGAGAAGTG, wzy-R:
81
GCAATCGAAGTGAAGAGTGC and rmpA-F: ACTGGGCTACCTCTGCTTCA, rmpA-
82
R: CTTGCATGAGCCATCTTTCA. Both hypermucoviscocity associated genes were
83
detected in the studied isolate.
84
Although K. pneumoniae ST23 isolates usually display a susceptible profile, here we
85
detected the presenceof KPC-2 in an isolate belonging to this invasive sequencetype. The
86
presence of KPC carbapenemases in K. pneumoniae ST23 has been only previously
87
reported in isolates from China and Poland, in 2010 and 2011, respectively, displaying the
88
same susceptibility profile as K. pneumoniae 3089 (17, 19, 20). However, no single
89
mention to the virulence factors or hypermucoviscosity phenotype was included in those
90
studies. 4
91
In the last few months three more hypermucoviscous K. pneumoniae ST23 isolates have
92
been referred to our laboratory, displaying antimicrobial susceptible phenotypes except for
93
ampicillin. Considering the virulence factors associated to this phenotype and its highly
94
invasive feature a prompt identification and accurate treatment should be mandatory. These
95
strains can be readily detected by the string test, MLST and molecular characterization of
96
the hypermucoviscous phenotype associated genes. Antibiotics commonly used in K.
97
pneumoniae infections have been useful for the therapeutic treatment of ST23 clinical
98
isolates, however the acquisition of resistance genes in this invasive strains could hinder
99
their eradication, probably favoring the development of metastatic infections.
100
A rising number of cases of K. pneumoniae ST23 from geographic regions other than
101
Southeast Asia indicate that this is a globally emerging pathogen. According to Brisse et
102
al., K. pneumoniae ST23 constitutes an emerging highly virulent and metabolically
103
versatile clone (14), so the acquisition of an important mechanism of antibiotic resistance
104
such as KPC-2 could mark an evolutionary step towards the settling of K. pneumoniae
105
ST23 as a major cause of nosocomial infections.
106 107
Acknowledgments
108
This work was partially supported by Grants from UBACyT and ANPCyT to M. Radice
109
and G. Gutkind. G. Gutkind and M. Radice are members of Carrera del Investigador
110
Científico (CONICET). D. Cejas was recipientof a doctoral fellowship from CONICET and
111
is now recipient of a post doctoral fellowship from Fundación Bunge y Born.
112 113 114 5
115
References
116
1.
117
antimicrobial susceptibility testing; twenty-first informational supplement M100-S23.
118
2013;33 (1).
119
2.
120
Brooks S, Cebular S, Quale J. 2004. Emergence of carbapenem-resistant Klebsiella
121
species possessing the class A carbapenem-hydrolyzing KPC-2 and inhibitor-resistant
122
TEM-30 beta-lactamases in New York City. Clin Infect Dis. 39:55-60.
123
3.
124
Genetic structures at the origin of acquisition of the beta-lactamase blaKPC gene.
125
Antimicrob Agents Chemother. 52:1257-1263.
126
4.
127
Identification of plasmids by PCR-based replicon typing. J Microbiol Methods. 63:219-
128
228.
129
5.
130
Lastovetska O, Albornoz E, Galas M, Melano RG, Corso A, Petroni A. 2011. Clonal
131
dissemination of Klebsiella pneumoniae ST258 harbouring KPC-2 in Argentina. Clin
132
Microbiol Infect. 17:1520-1524.
133
6.
134
carbapenemase-producing bacteria. Lancet Infect Dis. 9:228-236.
135
7.
136
Vay C, Maldonado I, Famiglietti A, Giovanakis M, Magarinos F, Berardinelli E,
137
Neira L, Mollerach M, Gutkind G, Radice M. 2012. Hyperendemic clone of KPC
Clinical and Laboratory Standards Institute. Performance standards for
Bradford PA, Bratu S, Urban C, Visalli M, Mariano N, Landman D, Rahal, JJ,
Naas T, Cuzon G, Villegas MV, Lartigue MF, Quinn JP, Nordmann P. 2008.
Carattoli A, Bertini A, Villa L, Falbo V, Hopkins KL, Threlfall EJ. 2005.
Gomez SA, Pasteran FG, Faccone D, Tijet N, Rapoport M, Lucero C,
Nordmann P, Cuzon G, Naas T. 2009. The real threat of Klebsiella pneumoniae
Cejas D, Fernandez Canigia L, Nastro M, Rodriguez C, Tanco A, Rodriguez H,
6
138
producing Klebsiella pneumoniae ST 258 in Buenos Aires hospitals. Infect Genet Evol.
139
12:499-501
140
8.
141
Faccone D, Di Martino A, Galas M. 2008. Klebsiella pneumoniae Carbapenemase-2,
142
Buenos Aires, Argentina. Emerg Infect Dis. 14:1178-1180.
143
9.
144
sequence typing of Klebsiella pneumoniae nosocomial isolates. J Clin Microbiol. 43:4178-
145
4182.
146
10.
147
with septic endophthalmitis. Arch Intern Med. 146:1913-1916.
148
11.
149
endemic disease, Taiwan. Emerg Infect Dis. 14:1592-1600.
150
12.
151
of colanic acid synthesis in Escherichia coli by rmpA and rmpB, two virulence-plasmid
152
genes of Klebsiella pneumoniae. Mol Microbiol. 3:1349-1359.
153
13.
154
the putative virulence gene magA in a worldwide collection of 495 Klebsiella isolates:
155
magA is restricted to the gene cluster of Klebsiella pneumoniae capsule serotype K1. J Med
156
Microbiol. 54:1111-1113.
157
14.
158
Grimont P. 2009. Virulent clones of Klebsiella pneumoniae: identification and
159
evolutionary scenario based on genomic and phenotypic characterization. PLoS
160
One.4:e4982.
Pasteran FG, Otaegui L, Guerriero L, Radice G, Maggiora R, Rapoport M,
Diancourt L, Passet V, Verhoef J, Grimont PA, Brisse S. 2005. Multilocus
Liu YC, Cheng DL, Lin CL. 1986. Klebsiella pneumoniae liver abscess associated
Tsai FC, Huang YT, Chang LY, Wang JT. 2008. Pyogenic liver abscess as
Nassif X, Honore N, Vasselon T, Cole ST, Sansonetti PJ. 1989. Positive control
Struve C, Bojer M, Nielsen EM, Hansen DS, Krogfelt KA. 2005. Investigation of
Brisse S, Fevre C, Passet V, Issenhuth-Jeanjean S, Tournebize R, Diancourt L,
7
161
15.
Turton JF, Englender H, Gabriel SN, Turton SE, Kaufmann ME, Pitt TL.
162
2007. Genetically similar isolates of Klebsiella pneumoniae serotype K1 causing liver
163
abscesses in three continents. J Med Microbiol. 56:593-597.
164
16.
165
Appearance of Klebsiella pneumoniae liver abscess syndrome in Argentina: case report and
166
review of molecular mechanisms of pathogenesis. Open Microbiol J. 5:107-113.
167
17.
168
plasmid carrying blaKPC-2 in Enterobacteriaceae isolates from distinct cities in China.
169
Microb Drug Resist. 16:297-301.
170
18.
171
pneumoniae in North America. J Clin Microbiol. 43:991-992.
172
19.
173
D, Kania-Pudlo M, Mlynarczyk G, Zak-Pulawska Z, Hryniewicz W, Gniadkowski M.
174
2011. Molecular characteristics of KPC-producing Enterobacteriaceae at the early stage of
175
their dissemination in Poland, 2008-2009. Antimicrob Agents Chemother. 55:5493-5499.
176
20.
177
producing Klebsiella pneumoniae in China. J Antimicrob Chemother. 66:307-312.
Vila A, Cassata A, Pagella H, Amadio C, Yeh KM, Chang FY, Siu LK. 2011.
Qi Y, Wei Z, Li L, Ji S, Du X, Shen P, Yu Y. 2010. Detection of a common
Fang FC, Sandler N, Libby SJ. 2005. Liver abscess caused by magA+ Klebsiella
Baraniak A, Grabowska A, Izdebski R, Fiett J, Herda M, Bojarska K, Zabicka
Qi Y, Wei Z, Ji S, Du X, Shen P, Yu Y. 2011. ST11, the dominant clone of KPC-
178
8
179
Figure 1: Genetic context of blaKPC-2.
180
Legend figure 1: Primers (5’-3’) used to perform the PCR mapping of blaKPC-2. KPC-F:
181
ATGTCAC TGTATCGCCGTCT, KPC-R:TTTTCAGAGCCTTACTGCCC (2), 816U:
182
CACCTACACCACGACGAACC, 3098U: TGACCCTGAGCGGCGAAAGC and 4714:
183
GAAGATGCCAAGGTCAATGC (3) and IstB-RV:TTCCTGACCACTCCCGCCTTCC
184
(this study).
185 186
Figure 2: Hypermucoviscous phenotype of K. pneumoniae 3089
187
Legend figure 2: Hypermucoviscous phenotype is characterized by the formation of
188
elongated (≥5 mm) mucoviscous strings when a loop is passed through a colony (positive
189
string test).
9
1kb
t npA
I s t A 816U
I s t B
bl aKPC-2
3098U
t npA
KPCFW
I s t BRV
KPCRV
4714
