An Investigation into Laboratory Misidentification of a Bloodstream Klebsiellavariicola Infection

Berry, Gregory J.; Loeffelholz, Michael J.; Williams‐Bouyer, Natalie

doi:10.1128/jcm.00841-15

Cited by 39 publications

(50 citation statements)

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“…variicola is a nitrogen-fixing bacterium that was first identified in 2004 based on phylogenetic analysis of the rpoB gene sequence and DNA-DNA hybridization. It is estimated that 2.5% to 10% of K. pneumoniae isolates are actual misidentifications of K. variicola (1)(2)(3)(4). K. variicola is likely under-recognized because of its similar phylogenetic and biochemical properties to those of K. pneumoniae.…”

Section: Discussionmentioning

confidence: 99%

“…variicola in the database and the isolate was misidentified as K. pneumoniae. The microbiology laboratory personnel suggested that we explore the possibility that our isolate was K. variicola based on a recent report by Berry et al on the misidentification of K. pneumoniae (1).…”

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confidence: 99%

“…DNA was extracted from the culture, and the KVAR_0717 gene of the Klebsiella species was PCR amplified and then subjected to Sanger sequencing. This gene is a target based on sequencing differences between K. variicola and K. pneumoniae (1). Based on sequence homology determined using the Basic Local Alignment Search Tool (BLAST) (https:// blast.ncbi.nlm.nih.gov/Blast.cgi), the isolate was identified as K. variicola.…”

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confidence: 99%

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The Brief Case: Klebsiella variicola —Identifying the Misidentified

et al. 2019

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A 72-year-old Japanese male with a history of colorectal cancer with metastases to the liver and peritoneum presented to the emergency department with fever and abdominal pain and was subsequently found to have Gram-negative bacteremia. The patient had undergone numerous chemotherapy treatment courses since his diagnosis 18 months prior as well as a central hepatectomy with hepaticojejunostomy. One year after the resection, he developed recurrent ascending cholangitis from a biliary stricture with an associated bile leak, necessitating the placement of a biliary stent and percutaneous drain. Despite drain placement, he presented to the hospital several times with Gram-negative bacteremia, most frequently growing either Escherichia coli or Klebsiella pneumoniae. Prior to transfer of his care to our institution 1 month prior, he had been placed on rotating prophylactic antibiotics, including amoxicillin-clavulanic acid, ciprofloxacin, and trimethoprim-sulfamethoxazole. Vital signs on admission revealed a fever of 103.0°F, heart rate of 100 bpm, and a blood pressure of 96/64 mm Hg. On exam he was cachectic, lethargic, and diaphoretic. His respiratory exam was notable for rales in the left lower base, and he had rightupper-quadrant abdominal pain with deep palpation but did not have rebound tenderness. His biliary drain contained scant amounts of dark green bilious fluid, and the rest of his exam was unremarkable. The initial complete metabolic panel (CMP) revealed an elevated alkaline phosphatase of 763 U/liter, which was above his baseline of 200 to 300 U/liter. His white blood cell (WBC) count was 6.99 ϫ 10 3 cells/mm 3. A paracentesis was performed which was inconsistent with spontaneous bacterial peritonitis, noting only 11/mm 3 neutrophils. A computed tomography (CT) scan of his abdomen and pelvis with contrast revealed that the percutaneous biliary drain terminated in the hepaticojejunostomy and that the intrahepatic bile duct remained undrained, raising concern for possible obstruction. Two sets of bioMérieux BacT/Alert FN Anaerobic Plus and FA Aerobic Plus blood cultures were obtained. One of the two sets grew Gram-negative bacilli that could not be identified by our lab's Verigene Gram-negative blood culture panel. The sample was next tested using a Biofire Filmarray blood culture identification panel, which returned K. pneumoniae. The blood cultures were sent to our reference lab, Kaiser Permanente Northwest Regional Laboratory, which identified the pathogen as K. pneumoniae using a Vitek mass spectrometry matrix-assisted laser desorption/ionization time of flight (MALDI-TOF MS) instrument with version 2.0 software. An adonitol fermentation test was also performed on a Vitek 2 instrument running version 7.01 software, which was negative. These discordant results suggested that this could be a Klebsiella species other than K. pneumoniae. Our institution's probe for K. pneumoniae used with the Verigene panel does not cross-react with Klebsiella variicola and therefore returned the result "no molecular ID det...

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Section: Discussionmentioning

confidence: 99%

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The Brief Case: Klebsiella variicola —Identifying the Misidentified

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“…Furthermore, a reliable ID may have been compromised by the quality of the specimen or due to the few differences in the proteomic profiles within each group in those cases where no spectral profile or no reliable ID was attained. Lastly, as noted in previous studies, the discrepancies observed in multiple blood culture samples (BC04, BC157, BC160, and BC192) were probably due to the MALDI-TOF MS and Vitek inability to differentiate between K. pneumoniae and K. variicola due to similar biochemical and proteomic profiles (17,22,23). With regard to the BC-GN, it seems more likely that this assay does not misidentify K. variicola isolates and correctly reports them as not identified since the BC-GN K. pneumoniae probe has a low enough sequence identity with K. variicola (88% versus 100% for K. pneumoniae) to select against K. variicola as a target (22,23).…”

Section: Discussionmentioning

confidence: 53%

Parallel Evaluation of the MALDI Sepsityper and Verigene BC-GN Assays for Rapid Identification of Gram-Negative Bacilli from Positive Blood Cultures

Arroyo

Denys

2017

J Clin Microbiol

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Rapid identification of microorganisms from positive blood cultures has improved clinical management and antimicrobial stewardship. The advent of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has reduced the time to identification of cultured isolates and is now often the definitive method used in the clinical microbiology laboratory. The commercial diagnostic MALDI Sepsityper (Sepsityper) kit has the potential for standardization and clinical routine use for the rapid identification of a broad range of bacteria from positive blood cultures. In this study, we performed a parallel evaluation of the Sepsityper (Bruker Daltonics, Billerica, MA) and the Verigene BC-GN (BC-GN) assays (Nanosphere, Inc., Northfield, IL) for the identification of Gram-negative bacilli. A total of 210 Bactec bottles demonstrating Gram-negative bacilli were prospectively enrolled for this study. Among these, 200 monomicrobial cultures were included in the comparative analysis. For monomicrobial cultures, the BC-GN detected 85% (170/200) compared to that detected by routine culture while the Sepsityper detected 94% (188/200) and 91% (181/200) to the genus and species levels, respectively. Comparable positive percentage agreement and negative percentage agreement were observed between the Sepsityper (96.5% and 98.8%, respectively) and the BC-GN (99.4% and 99.8%, respectively) when only ( = 170, 85%) organisms targeted by the latter test were included in the analysis. In conclusion, the two methods evaluated in this study showed excellent performance characteristics for the identification of Gram-negative bacilli commonly isolated from blood cultures. The Sepsityper showed a broader identification range capability that may further improve clinical management and antimicrobial stewardship in patients with less frequent Gram-negative bacilli bloodstream infections.

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“…In the case of the K. pneumoniae complex, MALDI-TOF MS identification remains largely unsatisfactory given the absence of well characterized, representative members of the complex in spectral databases. Currently, only K. pneumoniae and K. variicola are included in the Bruker database (https://www.bruker.com/fileadmin/user_upload/1-Products/Separations_MassSpectrometry/MALDI_Biotyper/US_CA_System/MBT_list_of_organisms10_2017.pdf), and identification of even these two species is imprecise given the lack of reference spectra of other phylogroups (13, 19). To address this important limitation of currently MALDI-TOF MS technology, we used a collection of well characterized reference strains from the six K. pneumoniae complex phylogroups and analyzed them by MALDI-TOF MS in order to define the potential of this method to identify species within the K. pneumoniae complex.…”

Section: Introductionmentioning

confidence: 99%

Identification of Klebsiella pneumoniae complex members using MALDI-TOF mass spectrometry

Rodrigues¹,

Passet²,

Brisse³

2018

Preprint

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29Klebsiella pneumoniae (phylogroup Kp1), one of the most problematic pathogens associated 30 with antibiotic resistance worldwide, is phylogenetically closely related to K. quasipneumoniae 31 [subsp. quasipneumoniae (Kp2) and subsp. similipneumoniae (Kp4)], K. variicola (Kp3) and two 32 unnamed phylogroups (Kp5 and Kp6). Together, Kp1 to Kp6 make-up the K. pneumoniae complex. 33Currently, the phylogroups can be reliably identified only by gene sequencing. Misidentification using 34 standard methods is common and the clinical significance of K. pneumoniae complex members is 35 therefore imprecisely defined. Here, we evaluated the potential of MALDI-TOF mass spectrometry to 36 discriminate K. pneumoniae complex members. We report for the first time the existence of mass 37 spectrometry biomarkers associated with the phylogroups, with a sensitivity and specificity ranging 38 between 80-100% and 97-100%, respectively. Strains within phylogroups Kp1, Kp2, Kp4 and Kp5 39 each shared two specific peaks not observed in other phylogroups. Kp3 strains shared a peak that was 40 only observed otherwise in Kp5. Finally, Kp6 had a diagnostic peak shared only with Kp1. Kp3 and 41Kp6 could therefore be identified by exclusion criteria (lacking Kp5 and Kp1-specific peaks, 42 respectively). Further, ranked Pearson correlation clustering of spectra grouped strains according to 43 their phylogroup. These results call for incorporation of spectra of all K. pneumoniae complex 44 members into reference MALDI-TOF spectra databases, in which they are currently lacking. This 45 advance may allow for simple and precise identification of K. pneumoniae and closely related species, 46 opening the way to a better understanding of their epidemiology, ecology and pathogenesis. 47 *

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An Investigation into Laboratory Misidentification of a Bloodstream Klebsiellavariicola Infection

Cited by 39 publications

References 8 publications

The Brief Case: Klebsiella variicola —Identifying the Misidentified

The Brief Case: Klebsiella variicola —Identifying the Misidentified

Parallel Evaluation of the MALDI Sepsityper and Verigene BC-GN Assays for Rapid Identification of Gram-Negative Bacilli from Positive Blood Cultures

Identification of Klebsiella pneumoniae complex members using MALDI-TOF mass spectrometry

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