JCM Accepted Manuscript Posted Online 3 June 2020

J. Clin. Microbiol. doi:10.1128/JCM.00582-20

Copyright © 2020 American Society for Microbiology. All Rights Reserved.

1 Newly-named Klebsiella aerogenes (formerly Enterobacter aerogenes) is

2 Associated with Poor Clinical Outcomes Relative to other Enterobacter Species in

3 Patients with Bloodstream Infection

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5 Authors: Austin Wesevich, MD, MPH1,2; Granger Sutton, PhD3; Felicia Ruffin, MSN4,

6 Lawrence P. Park, PhD4; Derrick Fouts, PhD3; Vance G. Fowler, Jr., MD, MHS4*; and

7 Joshua T. Thaden, MD, PhD4

9 1
Department of Medicine, Duke University School of Medicine, Durham, NC, USA

10 2
Department of Pediatrics, Duke University School of Medicine, Durham, NC, USA

11 3
J Craig Venter Institute, Rockville, MD, USA

12 4
Division of Infectious Diseases, Duke University School of Medicine, Durham, NC, USA

13

14 Keywords: Gram-negative bacteremia, Klebsiella aerogenes, Enterobacter cloacae

15 complex, bloodstream infection

16

17 Intended target: Original article

18

19 *Corresponding author:

20 Vance G. Fowler, Jr. MD, MHS

21 Division of Infectious Diseases, Duke University School of Medicine

22 Room 185 Hanes Building, 315 Trent Drive

23 Durham, NC 27710

24 Phone: +1-919-668-6053; Fax: +1-919-684-8902; E-mail: Vance.Fowler@duke.edu

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25 ABSTRACT

26 Objectives: Enterobacter aerogenes was recently renamed Klebsiella aerogenes. This

27 study aimed to identify differences in clinical characteristics, outcomes, and bacterial

28 genetics among patients with K. aerogenes versus Enterobacter species bloodstream

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29 infections (BSI).

30 Methods: We prospectively enrolled patients with K. aerogenes or Enterobacter cloacae

31 complex (Ecc) BSI from 2002-2015. We performed whole genome sequencing (WGS)

32 and pan-genome analysis on all bacteria.

33 Results: Overall, 150 patients with K. aerogenes (46/150 [31%]) or Ecc (104/150 [69%])

34 BSI were enrolled. The two groups had similar baseline characteristics. Neither total in-

35 hospital mortality (13/46 [28%] versus 22/104 [21%]; p=0.3) nor attributable in-hospital

36 mortality (9/46 [20%] versus 13/104 [12%]; p=0.3) differed between patients with K.

37 aerogenes versus Ecc BSI, respectively. However, poor clinical outcome (death before

38 discharge, recurrent BSI, and/or BSI complication) was higher for K. aerogenes than Ecc

39 BSI (32/46 [70%] versus 42/104 [40%]; p=0.001). In a multivariable regression model, K.

40 aerogenes BSI, relative to Ecc BSI, was predictive of poor clinical outcome (odds ratio

41 3.3; 95% confidence interval 1.4-8.1; p=0.008). Pan-genome analysis revealed 983

42 genes in 323 genomic islands unique to K. aerogenes isolates, including putative

43 virulence genes involved in iron acquisition (n=67), fimbriae/pili/flagella production

44 (n=117), and metal homeostasis (n=34). Antibiotic resistance was largely found in Ecc

45 lineage 1, which had a higher rate of multidrug resistant phenotype (23/54 [43%])

46 relative to all other bacterial isolates (23/96 [24%]; p=0.03).

47 Conclusions: K. aerogenes BSI was associated with poor clinical outcomes relative to

48 Ecc BSI. Putative virulence factors in K. aerogenes may account for these differences.

49

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50 INTRODUCTION

51 The Enterobacterales family of Gram-negative bacteria were originally divided into three

52 genera: Escherichia, Aerobacter, and Klebsiella, where the Aerobacter genus included

53 A. aerogenes and A. cloacae (1). By 1960, Aerobacter had been redubbed Enterobacter

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54 (2). Recently, whole genome sequence (WGS)-based comparative bacterial

55 phylogenetics demonstrated that Enterobacter aerogenes is more closely related to

56 Klebsiella pneumoniae than to the Enterobacter species (3). Hence, the bacteria

57 formerly known as Enterobacter aerogenes was renamed Klebsiella aerogenes (4). The

58 remaining members of the genus Enterobacter can be grouped into 22 distinct

59 phylogenetic groups using average nucleotide identity (ANI) (3, 5). These 22

60 phylogenetic groups are together referred to as Enterobacter cloacae complex (Ecc).

61

62 Though bacterial comparative genomics has demonstrated that K. aerogenes and Ecc

63 belong to different phylogenetic groups, the clinical impact of these genetic differences is

64 unknown. Differences in clinical risk factors, antibiotic susceptibility patterns, and patient

65 outcomes have not yet been explored since renaming K. aerogenes. Further, it is

66 unclear how genetic diversity within Ecc, which consists of 22 phylogenetic groups in

67 and of itself, influences antibiotic resistance and clinical outcomes. These questions are

68 relevant as we and others have shown that species-level differences in Gram-negative

69 bacteria infections are associated with differences in clinical risk factors (6, 7), antibiotic

70 susceptibility patterns (6, 8, 9), patient outcomes (6, 7, 10), and complications of

71 infection (11). Better understanding the clinical impact of different Gram-negative

72 bacterial infections allows for more appropriate empiric antibiotic therapy, improved

73 monitoring for infectious complications, and enhanced prognostic information.

74

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 75 The aims of this study are to identify how patients with K. aerogenes and Ecc

76 bacteremia differ with respect to patient characteristics, patient outcomes, bacterial gene

77 content, and antibiotic resistance phenotypes using prospectively collected clinical data

78 and bacterial isolates from a biorepository at Duke University Medical Center from 2002-

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79 2015. Patient clinical data was retrospectively analyzed, and bacterial whole genome

80 sequencing and pan-genome analysis were performed.

81

82 METHODS

83 Study Population

84 Patients in this study were identified from the Duke Bloodstream Infection Biorepository

85 (BSIB), which is an ongoing and prospectively enrolled cohort of patients with

86 monomicrobial Gram-negative bloodstream infection (BSI) at Duke University Medical

87 Center. Informed consent is obtained, and from each enrolled patient detailed clinical

88 data, bacterial isolate from blood culture, patient serum, and patient DNA are collected.

89 In this study, we included patients with K. aerogenes or Ecc BSI from 2002-2015. If

90 patients had multiple episodes of K. aerogenes or Ecc BSI, only the first such episode

91 was included. Patients were also excluded for age <18 years, polymicrobial BSI, or

92 culture drawn in outpatient setting.

93

94 Whole Genome Sequencing and Assembly

95 Whole genome sequencing was performed on the Enterobacterales isolates in

96 collaboration with J. Craig Venter Institute (JCVI). Genomic DNA was extracted with the

97 Master Pure DNA Purification kit (Epicentre) to construct paired-end libraries for whole

98 genome sequencing (Illumina HiSeq) with target coverage of 100X. HiSeq reads were

99 assembled using Newbler, velvet, and Celera Assembler, and annotation was performed

100 using NCBI's Prokaryotic Genome Annotation Pipeline (PGAP) (12).

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101

102 Genetic Analysis

103 Pan-genome analysis was performed on the Enterobacterales isolates in collaboration

104 with JCVI. Bioinformatics analyses including generation of a phylogenetic tree and

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105 identification of antibiotic resistance genes were performed through the JCVI pan-

106 genome pipeline (13). The Pan-genome Ortholog Clustering Tool (PanOCT) was used to

107 identify flexible genomic islands associated with K. aerogenes versus Ecc and patient

108 attributable mortality (13, 14). The PanOCT algorithm compared gene content using

109 default parameters except with a minimum percent identity set to 70%. PanOCT first

110 determines the core genes which are present in almost all of the isolates included in the

111 pan-genome (in this case both the K. aerogenes and Ecc isolates) and then determines

112 ‘flexible genomic islands’ (fgi) (15) that are inserted between core genes. These flexible

113 genomic islands and the genes they contain can then be queried for those that are

114 differentially present in K. aerogenes and Ecc BSI isolates.

115

116 Definitions

117 The BSI outcome was defined as one of four possible endpoints: 1) cure: no evidence of

118 recurrent infection within the hospital admission, 2) recurrence: clinical resolution of the

119 initial episode of infection after treatment but culture-confirmed BSI with the same

120 organism documented within the hospital admission, 3) attributable mortality: persistent

121 signs or symptoms of infection, positive blood cultures, or a persistent focus of infection

122 at the time of death in the absence of another explanation for death, or 4) death due to

123 underlying causes: deemed not to be due to K. aerogenes or Ecc BSI by evaluation of

124 one of the investigators (JTT). Complications including septic shock, acute kidney injury

125 (AKI), acute lung injury (ALI) or acute respiratory distress syndrome (ARDS),

126 disseminated intravascular coagulation (DIC), or shock liver were defined per standard

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127 guidelines (16-20). Poor clinical outcome was defined as presence of ≥1 of the following:

128 death prior to discharge, recurrent BSI, and/or complication of BSI. The multidrug

129 resistant (MDR), extensively drug-resistant (XDR), and pandrug resistant (PDR)

130 phenotypes were defined per standard guidelines: MDR as resistance to at least one

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131 drug in > 3 relevant classes, XDR as resistance to at least one drug in all but <2 relevant

132 classes, and PDR as resistance to all relevant drugs (21). Additional definitions may be

133 found in the supplemental material.

134

135 Statistical Analysis

136 Baseline characteristics and clinical events are presented as means with standard

137 deviation for continuous variables and frequencies with proportions for categorical

138 variables. Statistical comparisons between groups for continuous variables were made

139 with Mann-Whitney-U test or t-test. For categorical variables, comparisons were made

140 using Fisher’s exact test. Multivariable logistic regression models were used to

141 determine risk factors associated with total in-hospital mortality, attributable mortality,

142 and poor clinical outcome. Covariates in the multivariable logistic regression model

143 included BSI species, age, race, gender, route of BSI (i.e., community- vs. hospital-

144 acquired), source of BSI (e.g., genitourinary tract), days to effective antibiotic therapy,

145 and chronic APACHE-II score. Antibiotic resistance genes were compared using Fisher’s

146 exact test with Bonferroni correction for multiple comparisons. Analyses were done using

147 Stata 16.0.

148

149 RESULTS

150 Patient Demographics

151 There were 150 patients with K. aerogenes (46/150 [31%]) or Ecc (104/150 [69%]) BSI

152 enrolled over the study period. The clinical characteristics of patients with K. aerogenes

6
153 versus Ecc BSI were similar (Table 1). Patients with Ecc BSI were more commonly

154 hemodialysis-dependent (24/104 [23%]) compared to those with K. aerogenes BSI (4/46

155 [9%]; p=0.04). Otherwise, there were no significant differences between patients with K.

156 aerogenes versus Ecc BSI for rates of other clinical characteristics.

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157

158 Clinical Outcomes

159 The overall rates of total and attributable (i.e., due to the BSI) in-hospital mortality were

160 23% (35/150) and 15% (22/150), respectively. Neither total in-hospital mortality (28% vs.

161 21%, p=0.4) nor attributable in-hospital mortality (20% vs. 12%, p=0.3) differed

162 significantly between patients with K. aerogenes and Ecc BSI, respectively (Table 2).

163 The PanOCT algorithm did not identify any flexible genomic islands associated with

164 increased total or attributable in-hospital mortality in patients with K. aerogenes or Ecc

165 BSI. The overall rate of poor clinical outcome was 49% (74/150). Poor clinical outcome

166 was more common in patients with K. aerogenes BSI than Ecc BSI (70% vs. 40%,

167 p=0.001). The rates of all individual BSI complications were higher in K. aerogenes

168 versus Ecc BSI, though only acute kidney injury was statistically significant (43% vs.

169 21%, p=0.01). By definition, patients on hemodialysis cannot have the complication of

170 acute kidney injury. However, even when these 28 patients were excluded, those with K.

171 aerogenes BSI had increased rate of poor clinal outcomes (p=0.01). No bacterial clade

172 was associated with poor clinical outcome (Ecc, p=0.2; K. aerogenes p=0.6). Clinical

173 outcomes associated with the Ecc clades are shown in Table S1.

174

175 Multivariable logistic regression models for total in-hospital mortality, attributable

176 mortality, and poor clinical outcome were generated (Table 3). Bacterial species (i.e., K.

177 aerogenes versus Ecc) was not associated with either total in-hospital mortality (OR 1.2,

178 95% CI 0.4-3.0, p=0.8) or attributable mortality (OR 1.5, 95% CI 0.4-5.1, p=0.5). In the

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179 multivariable logistic regression model of poor clinical outcome, however, K. aerogenes

180 was associated with increased poor outcome (OR 3.3, 95% CI 1.4-8.1, p=0.008).

181

182 Bacterial Phylogeny

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183 Prior work demonstrated that there are 22 known Ecc clades (22). The 104 Ecc BSI

184 isolates included in this study encompassed 14 (64%) of these clades (Figure 1). More

185 broadly, the Ecc BSI isolates could be grouped into two lineages, with one lineage

186 encompassing Ecc clades A-E and the other encompassing clades G-J, L-M, O, and Q-

187 R (Figure 1). To our knowledge, K. aerogenes has no prior clade designations. Using an

188 average nucleotide identify approach, we identified three K. aerogenes clades, labeled

189 A-C (Figure 1). The K. aerogenes BSI isolates were primarily in the designated A clade

190 (39/45 [87%]).

191

192 The PanOCT algorithm was used to identify the specific genetic islands that distinguish

193 K. aerogenes from the Ecc BSI isolates. The total number of genes was 4392, with 3016

194 (69%) shared between K. aerogenes and Ecc, 983 (22%) unique to K. aerogenes, and

195 393 (9%) unique to Ecc (Figure 2). By specific to a given group we mean that 90% or

196 more of the isolates of that group contained the gene while 10% or fewer of isolates of

197 the other group contained the same gene. We identified 323 fgis specific to K.

198 aerogenes, and 191 specific to Ecc. Close inspection of genes unique to K. aerogenes

199 revealed multiple genes with putative functions in cellular processes that are often

200 associated with bacterial virulence (Table S2), including iron acquisition (8 fgi with 67

201 total genes), fimbriae/pili attachment (8 fgi with 106 total genes), flagella movement and

202 attachment (5 fgi with 11 total genes), and metal homeostasis (4 fgi with 34 total genes).

203

204 Antibiotic Resistance

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205 The rates of K. aerogenes and Ecc resistance to individual antibiotics are listed in Table

206 S3. Rates of resistance to trimethoprim/sulfamethoxazole (19% vs. 0%, p<0.001) and

207 gentamicin (10% vs. 0%, p=0.03) were higher in Ecc isolates relative to K. aerogenes.

208 The MDR phenotype was similar between Ecc and K. aerogenes (30% vs. 33%, p=0.8).

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209 Only Ecc had XDR isolates (3/104 [3%] versus 0/46 [0%]; p=0.6). No PDR isolates were

210 identified.

211

212 In order to determine the mechanisms by which K. aerogenes and Ecc develop

213 resistance to antibiotics, we used whole genome sequencing to identify antibiotic

214 resistance genes and antibiotic resistance mutations present in each bacterial isolate

215 (Figure 3). Penicillin binding protein 3 (PBP3) was the most commonly present

216 resistance gene, found in 94% (141/150) of K. aerogenes and Ecc isolates. PBP3 is an

217 essential protein that catalyzes cross-linking of the peptigoglycan wall, though the

218 variant in this study is homologous to PBP3 in Haemophilus influenzae, where it has

219 been implicated in resistance to β-lactam antibiotics.

220

221 Figure 3 illustrates increased acquisition of antibiotic genes within Ecc lineage 1 (i.e.,

222 clades A-E), which corresponds to species E. hormaechei (clades A-E).3 Ecc lineage 1

223 was associated with the MDR phenotype (23/54 [43%]) relative to Ecc lineage 2 (8/50

224 [16%]; p=0.005) and to all other non-Ecc lineage 1 bacterial isolates in this study (23/96

225 [24%]; p=0.03). The MDR phenotype in Ecc lineage 1 was driven by the acquisition of

226 multiple antibiotic resistance genes including the AAC/APH/ANT/AAD aminoglycoside

227 resistance genes (22/54 [41%]), ACT β-lactam resistance genes (33/54 [61%]),

228 TEM/SHV genes that confer the extended-spectrum β-lactamase phenotype (12/54

229 [22%]), gyrA/parC/parE fluroquinolone resistance genes (13/54 [24%]), and sul genes

230 that confer resistance to sulfonamides (18/54 [33%]) (Figure 3).

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231

232 DISCUSSION

233 This study aimed to understand differences between K. aerogenes and Ecc BSI with

234 respect to patient demographics, clinical outcomes, bacterial phylogenetics, and

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235 antibiotic susceptibility patterns. To our knowledge it is the first study to examine such

236 differences in the two patient populations. It revealed three major findings.

237

238 First, the clinical characteristics of patients with K. aerogenes versus Ecc BSI were

239 similar. Hemodialysis dependence was the only identified risk factor for Ecc BSI. Though

240 no prior studies have addressed the risk factors for BSI with K. aerogenes (or

241 Enterobacter aerogenes, as it was previously named) versus Ecc BSI, a large recent

242 study did find that hemodialysis dependence was an independent risk factor for

243 Enterobacter species BSI relative to non-infected control patients (23).

244

245 Second, we found that poor clinical outcome was significantly more common in patients

246 with K. aerogenes versus Ecc BSI. Poor clinical outcome is a collapsed variable that is

247 defined as present when the patient either dies prior to hospital discharge, has recurrent

248 infection, or experiences a complication of the BSI. A remarkable 70% of patients with K.

249 aerogenes BSI had poor clinical outcome. Patients with K. aerogenes BSI had

250 numerically higher rates of almost all the individual components that define this variable.

251 However, of these individual components only rate of AKI was statistically significant

252 between the two groups. Understanding why patients with K. aerogenes BSI had

253 increased rate of AKI is complicated by the fact that sepsis-induced AKI may occur

254 through diverse mechanisms. Sepsis-induced AKI is generally thought to stem from

255 global renal ischemia, cellular damage, and acute tubular necrosis (24). However, there

256 is a growing body of evidence to suggest that it may occur even in the absence of

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257 hypoperfusion. For example, cytokine levels (e.g., IL-6, IL-10) and oxidative stress have

258 been shown to also play roles in sepsis-induced AKI (25-27). While it is not clear why K.

259 aerogenes was associated with increased AKI, it is tempting to speculate that the

260 multiple additional iron acquisition genes may be playing a role as bacterial proliferation

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261 has been shown to be supplemented by iron sufficiency and suppressed by iron

262 restriction (28, 29). Further, elevated serum iron levels have been associated with

263 increased mortality in patients with sepsis (30).

264

265 Third, we found that antibiotic resistance was driven in large part by the acquisition of

266 resistance genes in Ecc lineage 1, which encompasses clades A-E. This lineage is E.

267 hormaechei (clades A-E). Ecc lineage 1 had significantly higher antibiotic resistance

268 relative to either Ecc lineage 2 or all other non-Ecc lineage 1 isolates included in this

269 study. As shown in Figure 3, this increased resistance stemmed from acquisition of

270 multiple genes that are active against aminoglycosides (AAC, APH, ANT, AAD), β-

271 lactams (ACT, TEM, SHV), fluoroquinolones (parC, parE, gyrA), and sulfonamides (sul

272 genes). The only XDR isolates (n=3) in this study were in Ecc lineage 1.

273

274 This study has several limitations. First, it involved only a single health system. However,

275 given that we enrolled a relatively large number of patients over a 14 year period, we

276 believe that our results are generalizable. Second, detailed management decisions are

277 not available for each patient to compare how treatment may have affected clinical

278 outcomes. However, we have included data involving timing of appropriate antibiotic

279 therapy, which is one of the most critical factors in sepsis outcomes.

280

281 In summary, the bacteria formerly known as Enterobacter aerogenes was recently

282 renamed Klebsiella aerogenes on the basis of whole genome sequence-based

11
283 phylogenetics. The clinical and bacterial virulence gene differences between K.

284 aerogenes and the remaining Enterobacter isolates had not been studied prior to this

285 work. Here we found that the clinical characteristics of patients in the two groups were

286 quite similar, though the clinical outcomes differed substantially. K. aerogenes BSI was

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287 associated with increased poor clinical outcome relative to Ecc BSI. The etiology behind

288 this difference is unknown, though could be related to the multiple putative virulence

289 genes (e.g., iron acquisition, flagella production, fimbriae production, metal homeostasis)

290 that were specifically identified in the K. aerogenes isolates.

291

292 Funding

293 Research reported in this publication was supported by the National Center for

294 Advancing Translational Sciences of the National Institutes of Health under Award

295 Number 1KL2TR002554 [J.T.T.] and NIH K24-AI093969 [V.G.F]. The content is solely

296 the responsibility of the authors and does not necessarily represent the official views of

297 the National Institutes of Health.

298

299 Acknowledgments

300 Portions of these results were previously presented in Washington, DC, USA at IDWeek

301 2019 (October 2-6, 2019; Poster #221).

302

303 Contribution

304 Authors contributions included conception and design (AW, GS, DF, VGF, JTT),

305 acquisition of data (FR, VGF, JTT), analysis and interpretation of data (AW, GS, DF, LP,

306 VGF, JTT), manuscript preparation (AW, GS, FR, LP, DF, VGF, JTT).

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403 25. Murugan R, Karajala-Subramanyam V, Lee M, Yende S, Kong L, Carter M,

404 Angus DC, Kellum JA, Genetic, Inflammatory Markers of Sepsis I. 2010. Acute

405 kidney injury in non-severe pneumonia is associated with an increased immune

406 response and lower survival. Kidney Int 77:527-35.

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407 26. Payen D, Lukaszewicz AC, Legrand M, Gayat E, Faivre V, Megarbane B,

408 Azoulay E, Fieux F, Charron D, Loiseau P, Busson M. 2012. A multicentre study

409 of acute kidney injury in severe sepsis and septic shock: association with

410 inflammatory phenotype and HLA genotype. PLoS One 7:e35838.

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411 27. Wu L, Gokden N, Mayeux PR. 2007. Evidence for the role of reactive nitrogen

412 species in polymicrobial sepsis-induced renal peritubular capillary dysfunction

413 and tubular injury. J Am Soc Nephrol 18:1807-15.

414 28. Freidank HM, Billing H, Wiedmann-Al-Ahmad M. 2001. Influence of iron

415 restriction on Chlamydia pneumoniae and C. trachomatis. J Med Microbiol

416 50:223-7.

417 29. Hsieh PF, Lin TL, Lee CZ, Tsai SF, Wang JT. 2008. Serum-induced iron-

418 acquisition systems and TonB contribute to virulence in Klebsiella pneumoniae

419 causing primary pyogenic liver abscess. J Infect Dis 197:1717-27.

420 30. Lan P, Pan KH, Wang SJ, Shi QC, Yu YX, Fu Y, Chen Y, Jiang Y, Hua XT, Zhou

421 JC, Yu YS. 2018. High Serum Iron level is Associated with Increased Mortality in

422 Patients with Sepsis. Sci Rep 8:11072.

423

424

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425 Figure legends

426

427 Figure 1. Phylogenetic tree of K. aerogenes and Enterobacter cloacae complex isolates

428 with clade designations. The number of bacterial isolates in each clade are listed.

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429

430 Figure 2. Venn diagram of genes shared and specific to Klebsiella aerogenes and

431 Enterobacter cloacae complex in bloodstream infection isolates.

432

433 Figure 3. Phylogenetic tree of K. aerogenes and E. cloacae complex (Ecc) with

434 presence of antibiotic resistance genes per strain. The colors in the inner ring indicate

435 the different Ecc clades (labeled A-E, G-J, L-M, O, Q-R) and K. aerogenes (all clades

436 same color). Antibiotic resistance genes in each strain are indicated by bars in the outer

437 concentric rings. The identified antibiotic resistance genes are noted in the key.

438

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439 Table 1. Demographics of patients with K. aerogenes and Enterobacter cloacae

440 complex (Ecc) bloodstream infections at Duke University from 2002-2015.

K. aerogenes Ecc
Clinical characteristics N=46 N=104 P-value
n (%) n (%)

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Age, median (IQR) 65 (59, 71) 57 (51, 68.5) 0.1
Race
White 28 (61) 72 (69) 0.2
Black 14 (30) 30 (29)
Other/unknown 4 (9) 2 (2)
Female gender 10 (22) 37 (36) 0.1
Past Medical History
Recent glucocorticoid use 12 (26) 19 (18) 0.3
Neoplasm 18 (39) 42 (40) 1.0
Diabetes mellitus 15 (33) 38 (37) 0.7
Transplant 5 (11) 12 (12) 1.0
Surgery in past 30 days 14 (30) 44 (42) 0.2
Hemodialysis dependence 4 (9) 24 (23) 0.04
Rheumatologic disorder 1 (2) 3 (3) 1.0
Site of acquisition
Community-acquired 21 (46) 57 (55) 0.4
Hospital-acquired 25 (54) 47 (45)
Source of infection
Urine/pyelonephritis 7 (15) 16 (16) 0.1
Pneumonia 4 (9) 4 (4)
Abscess 5 (11) 7 (7)
Line-associated 5 (11) 28 (27)
Skin/soft tissue 0 (0) 7 (7)
Biliary tract 5 (11) 6 (6)
Other 6 (13) 8 (8)
Source not identified 14 (30) 28 (27)
Days to appropriate antibiotic therapy, mean (SD) 0.8 (1.2) 0.7 (1.0) 0.4
Appropriate antibiotics on day 0 26 (57) 62 (60) 0.2
Appropriate antibiotics after day 0 19 (41) 41 (39)
APACHE-II acute physiology score, mean (SD) 8.6 (6.5) 7.1 (4.4) 0.1
APACHE-II chronic illness score, mean (SD) 3.9 (2.1) 3.8 (2.1) 0.8
Score of 0 10 (22) 25 (24) 0.8
Score of 5 36 (78) 79 (76)

441 SD = standard deviation; IQR = interquartile range

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442 Table 2. Outcomes of patients with K. aerogenes versus Enterobacter cloacae complex

443 (Ecc) bloodstream infections.

K. aerogenes Ecc
Outcomes N=46 N=104 P-value
n (%) n (%)
Total in-hospital mortality 13 (28) 22 (21) 0.4

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Attributable mortality 9 (20) 13 (12) 0.3
Recurrent bloodstream infection 0 (0) 2 (2) 1.0
Complications
Septic shock 14 (30) 22 (21) 0.2
Acute kidney injury 20 (43) 22 (21) 0.01
Acute lung injury/acute respiratory distress syndrome 4 (9) 6 (6) 0.5
Disseminated intravascular coagulation 2 (4) 1 (1) 0.2
Shock liver 1 (2) 2 (2) 1.0
Poor clinical outcome* 32 (70) 42 (40) 0.001
444 * ≥1 of the following: death prior to discharge, recurrent bloodstream infection, or
445 complication of bloodstream infection
446

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447 Table 3. Multivariable logistic regression models for total in-hospital mortality, attributable in-hospital mortality, and poor clinical

448 outcome in patients with K. aerogenes and E. cloacae complex bloodstream infection. Predictors with p<0.05 are in bold.

Predictor Total Mortality Attributable Mortality Poor Clinical Outcome

OR [95% CI] (p-value) OR [95% CI] (p-value) OR [95% CI] (p-value)
K. aerogenes 1.2 [0.4, 3.0] (0.8) 1.5 [0.4, 5.1] (0.5) 3.3 [1.4, 8.1] (0.008)
Age 1.0 [1.0, 1.1] (0.05) 1.1 [1.0, 1.1] (0.01) 1.0 [1.0, 1.1] (0.04)
Racea 0.7 [0.2, 1.8] (0.4) 0.3 [0.1, 1.2] (0.09) 0.7 [0.3, 1.6] (0.4)
Female gender 1.4 [0.5, 4.4] (0.5) 2.0 [0.5, 9.0] (0.3) 0.8 [0.3, 1.9] (0.6)
Hospital-acquired 1.1 [0.4, 3.0] (0.9) 2.6 [0.7, 9.8] (0.2) 1.8 [0.8, 4.3] (0.2)
Infection sourceb
Pneumonia 2.5 [0.2, 26.8] (0.4) 5.2 [0.2, 124.1] (0.3) 0.4 [0.05, 3.2] (0.4)
Abscessc 0.8 [0.1, 9.9] (0.8) - 0.2 [0.03, 1.0] (0.05)
Line 0.6 [0.1, 4.1] (0.6) 2.5 [0.2, 36.5] (0.5) 0.1 [0.03, 0.4] (0.002)
Skind - - 0.6 [0.1, 3.9] (0.6)
Biliary 1.7 [0.2, 13.5] (0.6) 7.3 [0.4, 119.5] (0.2) 0.4 [0.1, 1.9] (0.2)
Other 8.9 [1.4, 57.3] (0.02) 30.4 [2.0, 471.4] (0.02) 0.9 [0.2, 4.6] (0.9)
Unknown 6.4 [1.4, 29.8] (0.02) 8.1 [0.8, 88.4] (0.08) 0.6 [0.2, 2.0] (0.4)
Antibiotics after day 0 1.2 [0.4, 3.1] (0.7) 0.6 [0.2, 2.2] (0.5) 0.7 [0.3, 1.6] (0.4)
Chronic APACHE-II of 5 3.1 [1.0, 9.8] (0.06) 4.3 [0.9, 22.0] (0.08) 1.6 [0.01, 1.8] (0.1)
449 Abbreviations: OR, odds ratio; CI, confidence interval
450 a
Race analyzed as binary (white versus non-white).
451 b
Reference is patients with genitourinary source of infection
452 c
Abscess source not included in attributable mortality model due to small number of cases and events.
453 d
Skin source not included in total mortality and attributable mortality models due to small number of cases and events.

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