First time identification of Pandoraea sputorum from a patient with cystic fibrosis in Argentina: a case report
© The Author(s). 2017
Received: 13 July 2016
Accepted: 21 January 2017
Published: 7 February 2017
Pandoraea species are considered emerging pathogens in the context of cystic fibrosis (CF) and are difficult to identify by conventional biochemical methods. These multidrug resistant bacteria remain poorly understood particularly in terms of natural resistance, mechanisms of acquired resistance and impact on the prognosis of the disease and the lung function. Among them, Pandoraea sputorum has been previously described in few cases of CF patients from Spain, Australia, France and United States, underlining the need of more clinical data for a better knowledge of its pathogenicity. This is the first report relating to P. sputorum in a CF patient in Argentina.
Pandoraea sputorum was identified in a nine-year-old cystic fibrosis patient from Argentina, after treatment failure during an exacerbation. The isolates were successfully identified by combining molecular techniques based on 16S rRNA sequencing and mass spectrometry (MS) methods, after reassessing previous misidentified isolates by conventional methods. After first isolation of P. sputorum, patient’s clinical condition worsened but later improved after a change in the treatment. Although isolates showed susceptibility to trimethoprim–sulfamethoxazole and imipenem, in our case, the antibiotic treatment failed in the eradication of P. sputorum.
All combined data showed a chronic colonization with P. sputorum associated to a deterioration of lung function. We noted that the presence of P. sputorum can be underestimated in CF patients and MALDI-TOF MS appears to be a promising means of accurate identification of Pandoraea species.
KeywordsPandoraea sputorum Cystic fibrosis First report MALDI-TOF MS Argentina
In cystic fibrosis (CF) patients, the main cause of morbidity and mortality is the decline in the pulmonary function subsequent to pathogenic colonization with non-fermenting Gram negative bacteria (NFGNB) . Nevertheless, diverse bacterial species exhibit distinct degrees of pathogenicity and sensitivity patterns, requiring different clinical managements . Regularly, a series of opportunistic pathogens colonizes the CF lung throughout life, often culminating in chronic infection with Pseudomonas aeruginosa and, less frequently, with Burkholderia cepacia complex (Bcc) organisms. Other opportunistic Gram-negative bacteria are also encountered. They include various Enterobacteriaceae, Stenotrophomonas maltophilia, Achromobacter xylosoxidans [1, 3] and species that are only occasionally associated with human infections apart from CF, such as Cupriavidus, Ralstonia, and Pandoraea species. Some of these species have been associated with poor outcomes in CF, and the role that others may play in disease progression remains uncertain . Bacteria from the Pandoraea genus (P. pulmonicula, P. pnomenusa, P. apista, P. norimbergensis and P. sputorum) are NFGNB that are being considered as emerging multi-drug resistant pathogens in the context of CF [4, 5]. As they are still poorly known, particularly in terms of mechanisms of natural or acquired resistance and of their impact on lung function and the prognosis of the disease , more information is needed it to evaluate their pathogenic role. Conventional microbiology methods remain problematic for the accurate identification of Pandorea species and misidentification with bacteria of the genus Burkholderia or Ralstonia has been reported [6–8]. Moreover, isolates from CF patients with persistent pathogenic colonization often lose their characteristic phenotypes or growth conditions . Therefore, descriptions of infections due to Pandoraea species in CF patients could be underestimated. The availability of MALDI-TOF MS in clinical microbiology diagnosis allows a quick and better identification of bacterial species, especially Pandoraea species [5, 8, 10–12]. In this study we report for the first time in Argentina, a lung colonization with Pandoraea sputorum, in a young CF patient who did not respond to antibacterial empirical treatment. Identification was possible after reassessing historical identification results obtained by conventional methods, using MALDI-TOF MS and sequencing of 16S rRNA gene.
Antibiotic susceptibility testing was performed using a disk diffusion method according to the recommendations of the committee of CLSI . Susceptibility patterns were also identical for all isolates. In the absence of interpretive susceptibility criteria for strains of Pandoraea, results were interpreted using CLSI criteria for “other non-Enterobacteriaceae” isolates. The microorganism was resistant to amikacin, ceftazidime, cefepime, meropenem, gentamicin, aztreonam and colistin and susceptible to imipenem and trimethoprim-sulfamethoxazole. Minimum inhibitory concentrations (MIC, mg/L) were estimated by Vitek2 Compac® for amikacin (≥64), ceftazidime (≥64), cefepime (≥64), ciprofloxacine (≥4), colistin (≥16), meropenem (≥16), gentamicin (≥64), piperacillin-tazobactam (≥128), imipenem (≤0,25) and trimethoprim-sulfamethoxazole (TMS, ≤ 1/20), which confirms the susceptibility profile.
Discussion and Conclusions
In the last decade, many emerging multidrug resistant pathogens have been described in CF patients including new species of the Burkholderia cepacia complex, Brevundimonas diminuta, Inquilinus limosus, Acetobacter indonesiensis, Achromobacter xylosoxidans, Ochrobactrum anthropi, and bacteria of the genus Pandoraea [17, 18]. The pathogenicity of Pandoraea species remains controversial. These bacteria have been occasionally recovered from the lung transplant and/or respiratory tract of CF patients, as well as from blood cultures in non-CF patients and environmental samples [2, 4, 8, 19]. In our review of the bibliography, we found few cases of CF patients chronically colonized with P. sputorum: one case from Australia , two cases from Spain [10, 11], and one case from France . Interestingly, in our CF patient, S. aureus and P. aeruginosa were eventually isolated from respiratory tract cultures together with P. sputorum. This situation makes more difficult to know the real contribution of P. sputorum to the clinical condition of our patient. In this regard, intermittent or persistent colonization with Pandoraea species, that sometimes coincides with a deterioration in lung function, has been reported in addition to clinical evidence of invasive potential [8, 10, 11, 18]. Furthermore, besides the inability of Pandoraea species to produce a biofilm in the respiratory tract , it remains still unclear the role of different virulence factors, mechanisms of pathogenicity, the possibility of transmission between patients, and their role in lung damage in CF patients. This situation emphasizes the importance of having more data about clinical cases. The Pandoraea genus is multidrug resistant and treatment may be problematic. In our case, P. sputorum was only susceptible to imipenem and TMS, with a low MIC for imipenem (≤0,25 mg/L), and although the treatment failed to eradicate the bacteria, the clinical condition improved. This pattern of being resistant to carbapenem and meropenem but sensitive to imipenem, appears to be unique to most Pandoraea genus [5, 21], and it can be a practical guideline in the earliest identification steps.
It is documented, when applying only conventional identification phenotypic methods, the microbiology laboratory commonly misidentifies this pathogen as Ralstonia, Stenotrophomonas or Burkholderia species [6–8]. It has been also noted the limitations of the sequences of 16S rRNA and gyrB genes for differentiating the Pandoraea species [8, 22]. The accuracy and usefulness of MALDI-TOF as a routine technique for rapid identification of bacterium was demostrated. Nevertheless, one factor limiting the use of MALDITOF MS is the scant reference data sets for microorganisms that are infrequently isolated from clinical specimens . Due to these limitations, a polyphasic approach for an accurate identification is recommended. In our CF patient, all isolates were identified as P. sputorum, combining 16S rRNA PCR and sequencing, and a MALDI-TOF MS proteomic platform. Lastly, to determine persistence over time of the same P. sputorum isolate, as generally occurs with P. aeruginosa and Bcc in CF patients, a molecular epidemiological analysis was performed [2, 23]. BOX-PCR and ERIC-PCR showed indistinguishable patterns among all isolates of P. sputorum (includeding last isolate collected in April 2016), indicating a possible chronic colonization, which is defined here as three positive cultures of the same strain isolated within a 6-month period.
In conclusion, we noted that the presence of P. sputorum can be underestimated in CF patients when using conventional biochemical identification methods. We also showed that P. sputorum may have a potential role in pulmonary function deterioration when chronically colonized CF patients, emphasizing the need to implement control policies to protect CF community. This is particularly relevant, as in our case, the treatment failed despite sensitivity to imipenem and trimethoprim–sulfamethoxazole. Finally, MALDI-TOF MS appears to be an excellent tool for an accurate identification of organisms of this genus.
Clinical Laboratory Standards Institute
Forced expiratory volume in one second
- MALDI-TOF MS:
Matrix-assisted laser desorption ionization-time of flight mass spectrometry
Minimum inhibitory concentration
Non-fermenting Gram-negative bacteria
The authors are grateful to the staff of the microbiology labs at Hospital Pediátrico Dr. F. Barreyro for their work in supporting treatment of the child in this case. We also thank to Analia Fortte for proofreading the manuscript.
This work was partially funded by a grant from Fundación Roemmers (2014–2016).
Availability of data and materials
The datasets and materials during and/or analyzed during the current study available from the corresponding author on reasonable request.
PFM, JF, and MVS recognized the novelty of the case and drafted the manuscript. JF and MVS designed and coordinated the design and data acquisition. PFM, JF, and MVS analyzed and interpreted the results. JGF and ACB were responsible for treating the patients. PFM, LL, FA, MM, MB, CP, AB and AL were responsible for the microbiology analysis. MM drafted the manuscript. All authors read and approved the manuscript.
PFM and JF are researchers at the National University of Misiones and the National Scientific and Technical Research Council (CONICET).
The authors declare that they have no competing interests.
Consent for publication
Written informed consent was obtained from the parents of the patient for publication in this case report. A copy of the written consent is available for review by the Editor of this journal.
Ethics approval and consent to participate
Ethical approval to report this case was not required.
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