Catheter-Related Bloodstream Infection by Tsukamurella inchonensis in an Immunocompromised Patient

Abstract

We report a case of catheter-related bloodstream infection by Tsukamurella inchonensis, identified using 16S rRNA gene sequencing, in a patient with myelofibrosis who underwent a bone marrow transplant. Tsukamurella species infections are rare. To our knowledge, this is the first case of T. inchonensis bloodstream infection in an immunocompromised patient.

CASE REPORT

A 56-year-old woman was admitted to our hospital for a bone marrow transplant. She was diagnosed with primary myelofibrosis as the cause of pancytopenia 4 years earlier and had received metenolone and prednisolone as treatment. After a central venous catheter was implanted, she underwent a conditioning regimen of fludarabine and busulfan and, subsequently, a bone marrow transplant from an unrelated donor. Methotrexate and tacrolimus were administered to prevent graft-versus-host disease. Ten days posttransplant, she developed a fever (37.9°C), but radiography did not identify any focus of infection. The white blood cell count was 0.1 × 10⁹ cells/liter, and the C-reactive protein level was 90 mg/liter. Empirical treatment with cefepime (4 g/day) was initiated. After 3 days, her fever rose to 38.3°C, and cefepime was therefore discontinued and replaced by meropenem (3 g/day). Gram-positive rods, which were unidentifiable by routine microbiological examinations, were detected in the blood cultures. Upon treatment, the patient became afebrile, the C-reactive protein level decreased to 2.1 mg/liter, and the white blood cell count recovered to 1.7 × 10⁹ cells/liter 24 days posttransplant. On day 33 posttransplant, the fever recurred. Blood cultures were collected and the central venous catheter was removed. Identical, unidentifiable Gram-positive rods were detected both in the blood culture and on the catheter tip. Subsequently, the fever improved, and because the patient remained afebrile and the blood cultures came back negative, she was uneventfully discharged from the hospital.

As described above, the blood cultures were positive for Gram-positive rods in aerobic bottles of the Bactec 9240 system (BD, Franklin Lakes, NJ, USA) twice, after 92 h and 33 h of incubation. Gram staining of the specimens revealed straight Gram-positive rods (Fig. 1A). The specimens from the bottles were plated onto Trypticase soy agar II with 5% sheep blood (BD) and incubated at 35°C in ambient air. After 48 h, white, rough, dry, and nonhemolytic colonies, 2 mm in diameter, were observed (Fig. 1B). The colonies were found to be catalase positive and oxidase negative, and the isolates stained weakly acid-fast using the Kinyoun stain. The RapID CB Plus system (Remel Inc., Lenexa, KS, USA) was used for identification. The isolate was initially identified as Rhodococcus equi (microcode, 060751; probability level, 99.9%). However, the features of the colonies described above were clearly different from those of R. equi colonies, which are yellowish to salmon-pink or red, convex, and creamy with complete edges (1).

FIG 1.

(A) Gram staining of Tsukamurella inchonensis isolated from blood cultures (oil immersion, ×1,000); (B) colonial appearance of T. inchonensis on a sheep blood agar plate after 48 h of incubation.

Therefore, we performed molecular identification by PCR amplification and sequence analysis of the 16S rRNA gene using DNA extracted from the isolates. The universal primers 8UA (5′-AGAGTTTGATCMTGGCTCAG-3′) and 1485B (5′-ACGGGCGGTGTGTRC-3′) were used as described previously (2). BLAST searches of the GenBank entries were used for sequence analysis, and CLUSTAL W (neighbor-joining method) was used for sequence editing and phylogenetic analysis using TreeView. The 16S rRNA gene (accession number AB907635) showed a 99.9% sequence identity (1,439 bp over the entire 1,441-bp fragment) to that of Tsukamurella inchonensis (DSM 44067^T; accession number X85955). Therefore, we identified the isolate as T. inchonensis.

The biochemical features of the isolate were consistent with the reported features of the type strain (IMMIB D-771; same as DSM 44067^T and ATCC 700082) of T. inchonensis (3), except for its inability to utilize cellobiose as a carbon source. IMMIB D-771 has the ability to utilize cellobiose, whereas IMMIB AL-1155 (3) and our isolate did not. As the numbers of species within the aerobic actinomycetes grow, it seems that phenotypic characterization by biochemical testing is no longer sufficient to adequately characterize these species.

The antibiotic susceptibility of the isolate was determined after 20 h of incubation by the broth microdilution method using a commercially prepared microtiter plate containing a series of antimicrobials (RSMP2, RSCP2; Nissui Pharmaceutical, Tokyo, Japan). The MICs of the tested antimicrobials are shown in Table 1. The susceptibilities were interpreted by the criteria established by the Clinical and Laboratory Standards Institute (CLSI) (4), even though these criteria are provided primarily for Nocardia species.

TABLE 1.

MICs for Tsukamurella inchonensis isolated from the patient

Antimicrobial agent	MIC (μg/ml)	Susceptibilitya
Benzylpenicillinb	8	NA
Ampicillinb	16	NA
Amoxicillin-clavulanate	>8	NA
Cefotaxime	1	S
Cefaclorb	16	NA
Cefepime	1	S
Imipenem	0.25	S
Meropenemb	4	NA
Erythromycinb	≤0.125	NA
Clindamycinb	≤0.25	NA
Minocycline	≤0.25	S
Levofloxacinb	0.5	NA
Moxifloxacin	≤0.5	S
Vancomycinb	1	NA
Trimethoprim-sulfamethoxazole	>4	R

^aThe susceptibilities were interpreted according to the criteria developed by the Clinical and Laboratory Standards Institute (4). S, susceptible; R, resistant; NA, not assessed.

^bIndicates the antimicrobials that are not recommended by the CLSI.

Tsukamurella species are aerobic Gram-positive rods that are found in a broad range of environments such as soil, water, and sludge. The genus Tsukamurella, which currently contains 12 species with validly published names, belongs to the order Actinomycetales (1, 5), and T. inchonensis was reported as a new species of the genus on the basis of its 16S rRNA sequence and physiological characteristics (3). Because Tsukamurella species share many phenotypic characteristics with other species of the mycolic acid-containing genera of Actinomycetales, they can easily be misidentified when standard biochemical tests are used (5, 6). In fact, the isolate in this report was originally identified as Rhodococcus equi by the biochemical identification kit. However, as the morphology of the colonies was different from that of R. equi (1), we performed a sequence analysis of the 16S rRNA gene, leading to the positive identification of T. inchonensis.

Tsukamurella species have been very rarely reported as etiologic pathogens implicated in bloodstream or pulmonary infections in immunocompromised patients receiving chemotherapy, transplants, or hemodialysis. To date, only 21 cases of Tsukamurella species as the cause of bloodstream infections in immunocompromised patients have been reported (7–16), and these are summarized in Table 2. Similar to our case, 20 out of these 21 cases were catheter-related infections, with Tsukamurella pulmonis and Tsukamurella tyrosinosolvens accounting for the majority (62%) of cases. To our knowledge, this is the first reported case of T. inchonensis bloodstream infection in an immunocompromised patient. Kattar et al. reported that the ATCC 25938 strain of Tsukamurella paurometabola actually represented a misnamed T. inchonensis strain by 16S rRNA sequencing (6). In 4 T. paurometabola cases (7, 8), the isolates were identified not by sequencing but by biochemical features. Thus, it might be possible that some T. paurometabola cases were actually T. inchonensis cases. Although not in an immunocompromised patient, T. inchonensis bloodstream infection was previously reported in a patient who ingested hydrochloric acid (17), and this was also reported to be a central venous catheter-related infection. The type strain IMMIB D-771 originated from this isolate (3).

TABLE 2.

Previously reported cases of immunocompromised patients with bloodstream infections caused by Tsukamurella species

Tsukamurella sp.	No. of casesa					Reference(s)
	Hematological tumor		Solid tumor chemotherapy	HD	Transplant
	Chemotherapy	SCT
T. paurometabola	(1)		(2)	(1)		7, 8
T. pulmonis		2	1	2c	1	9, 15, 16
T. tyrosinosolvens	1	1	4b	1		9, 10, 11, 13, 14
T. strandjordae		1				9
T. inchonensis		1				This case
Not typed	1, (1)	1				9, 12

^aSCT, stem cell transplant; HD, hemodialysis. Parentheses indicate the cases in which 16S rRNA sequencing was not performed for identification.

^bOne out of the 4 cases was not a catheter-related infection (13).

^cOne out of the 2 cases recovered without catheter removal (15).

The antibiotic susceptibility properties of T. inchonensis have not yet been established. According to a case reported by Chong et al. (17) and this case, T. inchonensis showed susceptibility to some of the third-generation and extended-spectrum cephalosporins, fluoroquinolones, macrolides, tetracyclines, and imipenem and resistance to penicillins. Other species of Tsukamurella have similarly been found to be susceptible to fluoroquinolones and imipenem but resistant to penicillins and cephems (9, 12–14). T. tyrosinosolvens is susceptible to macrolides (9, 13), whereas other species conversely show resistance to this drug class (9, 12, 14). In 16 out of 17 cases where the removal of the venous catheter was reported, the removal was required for the patient's recovery, suggesting that removing the catheter is probably the most effective management strategy for these patients.

In conclusion, Tsukamurella species, including T. inchonensis, should be considered the pathogenic cause of catheter-related bloodstream infections, especially during the presence of unidentifiable Gram-positive rods in immunocompromised patients. Identification of the exact species by 16S rRNA gene sequence analysis is required, as the antibiotic susceptibilities differ among Tsukamurella species. Further analyses of similar cases are required to establish the most adequate diagnostic methods and treatment regimens for T. inchonensis infections.

Nucleotide sequence accession number.

The 16S rRNA gene from this case report was deposited in GenBank under accession number AB907635.