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Previous Article | Next Article 
Journal of Clinical Microbiology, February 2007, p. 641-644, Vol. 45, No. 2
0095-1137/07/$08.00+0 doi:10.1128/JCM.01938-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.
| CASE REPORT |
Department of Laboratory Medicine, The University of Texas M. D. Anderson Cancer Center, Houston, Texas
Received 18 September 2006/ Returned for modification 7 November 2006/ Accepted 14 November 2006
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ABSTRACT |
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CASE REPORT |
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The gram-negative organism did not ferment glucose and was characterized by (i) tan-yellow and dry colonies that wrinkled with time on 5% sheep blood agar, (ii) slow growth on MacConkey agar, (iii) cytochrome oxidase positivity, (iv) the ability to reduce nitrate, and (v) the ability to hydrolyze gelatin (Table 1). Identification with the Vitek GNI+ card and the API 20NE system (bioMérieux Inc., Durham, NC) was unsuccessful (API biotype 1010004, 86% Moraxella lacunata), so a 571-bp segment of the 16S rRNA gene was amplified and sequenced. Query of the GenBank database (NCBI) by use of the BLAST program (http://www.ncbi.nlm.nih.gov/BLAST) revealed 100% identity (571/571 bp) with a recently named species, Dokdonella koreensis (family Xanthomonadaceae; GenBank accession no. AY987368) (16). Antimicrobial susceptibility testing was performed with epsilometer strips (E-test; AB Biodisk, Solna, Sweden) on Mueller-Hinton agar (BBL, BD Microbiology Systems, Cockeysville, MD), according to the manufacturer's recommendations. MICs were relatively low for all agents tested, with the exception of amikacin (Table 2).
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Case 2. The patient was a 54-year-old man with a history of nodular sclerosing Hodgkin's disease. Midway through his third cycle of chemotherapy, the patient presented with fever (38.3°C), chills, and leukocytosis (15,000 leukocytes/µl; his normal baseline was 6,200 leukocytes/µl). He was admitted to the hospital, but no source of infection was readily identified. Blood and urine samples for culture were collected, and empirical antimicrobial therapy with vancomycin and cefepime was initiated. By use of the Isolator system, the growth of 200 to 500 CFU/10 ml blood of a coagulase-negative Staphylococcus sp. was detected from his CVC sample but not from a peripheral one. Additionally, a gram-negative bacillus was isolated from BACTEC bottles into which blood drawn both centrally (after 14 h of incubation) and peripherally (150 h) had been inoculated. The patient's CVC was removed.
The gram-negative organism was characterized by (i) an inability to ferment glucose, (ii) lavender-gray, wet colonies on blood and chocolate agar plates (BBL), (iii) cytochrome oxidase positivity, and (iv) nitrate reduction and (v) esculin hydrolysis capabilities (Table 1). Once again, neither the Vitek card nor the API 20NE system provided an acceptable identification (API biotype 141004), so amplification and sequencing of the near full-length 16S rRNA gene was performed. A query with the BLAST program revealed 99.6% identity (1,405/1,416 bp) with a recently named species, Aquimonas voraii (family Xanthomonadaceae; GenBank accession no. AY544768) (15). Antimicrobial susceptibility testing with epsilometer strips revealed generally low MICs, with the exception of those of trimethoprim-sulfamethoxazole (Table 2). The patient was discharged after 5 days and subsequently received an additional 10 doses of ceftriaxone therapy through a newly placed CVC. He has remained asymptomatic and blood culture negative since discharge.
Case 3. The patient was an 82-year-old man, referred to our institution for myelodysplastic syndrome, who had a nonhemolytic febrile transfusion reaction following the administration of six units of pooled platelets and two units of packed red blood cells through his CVC. His maximum temperature of 39.3°C defervesced within 2 h of antipyretic therapy; and he experienced no signs or symptoms of hypotension, dyspnea, or wheezing. Two sets of blood for quantitative cultures were drawn, and the remaining platelet product was sent for routine bacterial culture. The patient returned to his home state the next day with instructions to follow up if fever or other symptoms of infection appeared.
Culture of the pooled platelet product was sterile, yet two gram-negative bacilli were cultivated from a CVC blood culture, with each organism present in numbers exceeding 1,000 CFU/10 ml blood. One of the isolates was concurrently cultivated from the peripheral draw site (1 CFU/10 ml blood). The patient and his referring physician were notified of the preliminary report, whereupon he was admitted to his local, outlying hospital. Afebrile and without complaint at that time, the patient received empirical ceftriaxone (a single dose) as well as azithromycin and levofloxacin, based upon the presence of bilateral chest infiltrates. He was discharged 4 days later on levofloxacin without having his CVC removed. Subsequent blood cultures have been sterile, and symptoms of infection have not recurred.
The isolate present only in the CVC sample was identified as Pantoea agglomerans. The second gram-negative organism, present in CVC and peripheral blood, formed smooth, tan-yellow colonies and demonstrated (i) cytochrome oxidase positivity and (ii) gelatin hydrolysis capability but was unable to (iii) hydrolyze esculin, (iv) reduce nitrate, or (v) ferment glucose (Table 1). Because identification could not be rendered by the Vitek card or the API 20NE system (API biotype 0056104, 87% Chryseobacterium meningosepticum), amplification and sequencing of the 16S rRNA gene (to 1,478 bp) was performed. The nearest match to any validly named bacterial species by use of a search with BLAST was with three strains of Luteibacter rhizovicina (family Xanthomonadaceae), each of which shared 97.4% identity (1,398/1,434, 1,396/1,432, and 1,403/1,440 bp; GenBank accession nos. AJ580498, AJ580497, and AY785744, respectively) (10). Based upon the general consensus regarding bacterial identification by the method of 16S rRNA gene matches (1), this organism was considered a Luteibacter sp. Susceptibility testing with epsilometer strips demonstrated generally low MICs for all agents tested (Table 2).
The family Xanthomonadaceae was proposed in 2005 to contain 12 genera, including Stenotrophomonas, Xanthomonas, and other established genera (14). Since then, an additional 6 novel genera have been proposed, making a total of 18 genera in the family (http://www.bacterio.cict.fr/classifgenerafamilies.html; accessed 18 September 2006). Like other nonfermenting gram-negative bacilli, these organisms are ubiquitous in the environment and can be isolated from diverse sources in both aqueous and terreous habitats (6). Among the members of the family, S. maltophilia is the most prominent species in the medical literature and has been well described as a cause of significant nosocomial infections (3). Yet, little is known regarding the potential for human colonization and/or infection by related genera. Recent descriptions of three new species, Dokdonella koreensis, Aquimonas voraii, and Luteibacter rhizovicina, were each the result of cultivation from soil or water sources (10, 15, 16). Our cases occurred from December 2005 to June 2006 at The University of Texas M. D. Anderson Cancer Center, a 500-bed comprehensive center in Houston. Approximately 30,000 blood cultures were performed annually by using both the BACTEC 9240 automated culturing system (Plus Aerobic/F bottles; BD Diagnostic Systems, Sparks, MD) and the Isolator 10 system (Wampole Laboratories). When an Isolator tube culture was positive, the number of CFU was quantitated from the 10 ml of blood cultured. All subcultures were plated on blood agar, chocolate agar, and MacConkey agar (BBL, BD Microbiology Systems, Cockeysville, MD); and the cultures were incubated aerobically at 35°C with 5% CO2. Definitive identification of all organisms was reached through amplification of a central segment of the 16S rRNA gene (
590 bp) by using a set of universal primers (forward primer, 5'-TGCAGCCGCGGTAATAC-3'; reverse primer, 5'-CGCTCGTTGCGGGACTTAACC-3') (7). For the two organisms without full matches, nearly full-length segments of the gene were obtained through the use of additional primers (5'-GCGTGCTTAACACATGCAAGTC-3' and 5'-AGGAGGTGATCCAACCGCA-3') (7, 8).
It is clear that central venous catheterization is a risk factor for the development of bloodstream infections, particularly among oncology patients in whom such devices are maintained for prolonged periods (4). Given the presence of fever, all the cases presented here met the criteria for laboratory-confirmed bloodstream infection by use of a CDC-proposed definition (12). In case one, Dokdonella koreensis was isolated from blood drawn through a CVC on separate occasions 35 days apart, suggesting catheter colonization rather than actual bacteremia. Yet, the correlation with fever onset and the response to antimicrobial therapy support the notion of genuine infection. The two remaining cases illustrate documented catheter-related bloodstream infections caused by Aquimonas voraii and Luteibacter sp., based on differential times to positivity and differential colony counts, respectively. Both infections were complicated by the involvement of an additional bacterium from the CVC culture. Interestingly, case 3 was identified in the setting of an acute, nonhemolytic febrile transfusion reaction-related workup. It is known that the transfusion of blood products may pose an independent risk for the development of bacteremia (9) and that CVCs in place for a long time tend to be colonized intraluminally (13); we therefore wonder if transient or sustained bacteremia following product transfusion through such CVCs may account for some febrile, nonhemolytic transfusion reactions among cancer patients.
To the extent that they were analyzed, the isolates described here demonstrated different biochemical profiles compared with those of type strains of the species D. koreensis (16), A. voraii (15), and L. rhizovicina (10) (Table 1). It is unclear at present, due to the limited number of strains and the limited experience with these organisms, whether discrepancies reflect true phenotypic variability among species (as with A. voraii), the heterogeneity of different species within a single genus (as with Luteibacter), or simply variations in testing methodologies (as our results were generated largely from reactions with the API 20NE system). Even in the relative absence of biochemical reactivity, however, 16S rRNA gene sequencing provided sufficient taxonomic discrimination in all three cases.
Apart from cytochrome oxidase positivity and colony morphology, a major difference in the antimicrobial susceptibility patterns between the organisms reported here and S. maltophilia was noted. Whereas a high proportion of S. maltophilia strains exhibit resistance to aminoglycosides and many beta-lactams (3) (Table 2), we found generally low MICs for the three genera reported here. By using the breakpoints for nonmembers of the family Enterobacteriaceae, as outlined in Clinical and Laboratory Standards Institute (CLSI) guidelines (2), exceptions included resistance to amikacin by both Dokdonella isolates and to trimethoprim-sulfamethoxazole by the Aquimonas strain (Table 2). Prior antibiotic therapy has been demonstrated to be a risk factor for infection by S. maltophilia, probably by providing a selective advantage to inherently resistant organisms (5, 11). Two of the patients described here had received antimicrobial therapy prior to the development of symptomatic bloodstream infections: treatment with levofloxacin and ciprofloxacin preceded the initial and the subsequent Dokdonella isolations, respectively, by just a few days for case 1, and a course of amoxicillin-clavulanate was prescribed for upper respiratory symptoms 45 days before positive blood culture results for case 2. Based on (i) the in vitro susceptibilities to quinolone agents by both Dokdonella strains (Table 2) and (ii) the time interval between empirical therapy and Aquimonas bacteremia, however, we assume that selective pressure played a comparatively little role in predisposing the patients to infection by these organisms.
In summary, this report describes three opportunistic catheter-associated bloodstream infections caused by newly described organisms from the family Xanthomonadaceae. The application of rRNA gene sequencing for identification provided a more accurate taxonomic designation than traditional kit-based biochemical methods for these three isolates. Finally, given the ecological, microbiological, and pathogenic characteristics of S. maltophilia and related xanthomonads, we speculate that catheter colonization and subsequent infection among these three patients occurred after some unknown environmental exposure and that a selection advantage as a result of prior antimicrobial therapy probably played little role in the subsequent development of infection.
Nucleotide sequence accession number. The 16S rRNA gene sequence of the Luteibacter sp. from case patient 3 has been deposited in GenBank under accession number DQ986460.
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ACKNOWLEDGMENTS |
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We thank Brenda Snider for assistance, as well as Kathy Bernard and Gerry Saddler for helpful discussions.
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FOOTNOTES |
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Published ahead of print on 22 November 2006.
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REFERENCES |
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| Antimicrob. Agents Chemother. | Clin. Microbiol. Rev. |
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