Brevundimonas spp: Emerging global opportunistic pathogens

ABSTRACT Non-fermenting Gram-negative bacteria are problematic in clinical locations, being one of the most prevalent causes of nosocomial infections. Many of these non-fermenting Gram-negative bacteria are opportunistic pathogens that affect patients that are suffering with underlying medical conditions and diseases. Brevundimonas spp., in particular Brevundimonas diminuta and Brevundimonas vesicularis, are a genus of non-fermenting Gram-negative bacteria considered of minor clinical importance. Forty-nine separate instances of infection relating to Brevundimonas spp were found in the scientific literature along with two pseudo-infections. The majority of these instances were infection with Brevundimonas vesicularis (thirty-five cases – 71%). The major condition associated with Brevundimonas spp infection was bacteraemia with seventeen individual cases/outbreaks (35%). This review identified forty-nine examples of Brevundimonas spp. infections have been discussed in the literature. These findings indicate that infection review programs should consider investigation of possible Brevundimonas spp outbreaks if these bacteria are clinically isolated in more than one patient.


Introduction
Gram-negative, non-fermenting bacteria are an emerging concern in clinical locations, being a common cause of nosocomial infections. Opportunistic pathogens from this group include many different bacterial species, including: Acinetobacter baumannii, Burkholderia cepacia, Ralstonia pickettii, Pseudomonas aeruginosa, Sphingomonas pauciomobilis, and Stenotrophomonas maltophilia [1][2][3][4][5][6][7][8]. The group can survive in a wide variety of environments including different water sources (aircraft water, bottled water, hospital water, purified water) [9][10][11][12], and are usually resistant to a wide array of antimicrobials [13,14]. Examples include resistance to penicillins, aminoglycosides and monobactems in R. pickettii [13] and penicillins, aminoglycosides, carbapenems and monobactems in S. maltophilia [14]. Bacteria such as these have the ability to infect patients/individuals with underlying medical conditions and diseases. Examination of the scientific literature showed multiple types of infections resulting from Brevundimonas spp. This indicates that the genus may be a more widespread pathogen than was hitherto thought, with infections caused by Brevundimonas spp. being invasive and severe. The goal of this study was to give an overview of the range of Brevundimonas spp infections, any underlying conditions associated with Brevundimonas spp infections and the treatment options used in the treatment of any Brevundimonas spp infections in order to assist medical practitioners.

Genus Brevundimonas
The genus Brevundimonas was first proposed by Segers et al [15]; incorporating Pseudomonas diminuta and Pseudomonas vesicularis [16,17]. Several species of the genus Caulobacter were later transferred to Brevundimonas significantly emending the description of the genus [18]. Currently, there are 25 species with valid published names within the Brevundimonas genus (http://www.bac terio.net/brevundimonas.html). The type species is Brevundimonas diminuta; with the type strain being LMG 2089.

Brevundimonas diminuta
Brevundimonas diminuta is the type species of the Brevundimonas genus. It has been isolated from clinical specimens, including blood and urine [15] as well as from the lung sputum of cystic fibrosis patients [29]. B. diminuta is not believed to be a significant pathogen and its virulence is generally low. B. diminuta is used as a test organism to validate reverseosmosis (RO) filtration devices for drinking water purification and is also used to test the porosity of pharmaceutical-grade filters (0.2 mm) due to the small size of the bacterium when grown in minimal media [30,31]. The bacterium has however been shown to be capable of penetrating these filters [32]. The bacterium has been used as a potential bioremediator of marine oil pollution including diesels, n-alkanes and polycyclic aromatic hydrocarbons [33,34] and insecticides [35]. B. diminuta has also been used to mitigate the toxic effects of heavy metals on plant growth (rice) in contaminated soils [36]. B. diminuta also possesses the ability to survive sanitizers such as Hydrogen Peroxide + Peracetic Acid [37]. All available reported incidences of infection credited to B. diminuta are listed in Table 1-3.

Brevundimonas vesicularis
Brevundimonas vesicularis has been isolated from eye, urine, wound cultures, the central nervous system, cervical specimens [38], and also been found in the lung sputum of cystic fibrosis patients [39]. The organism has been shown to support the growth of Legionella in nutrient limited water conditions [40]. The mechanism behind this phenomenon has not been elucidated but it is hypothesised to be due to cryptic growth, with B. vesicularis having the ability to grow in nutrient limited conditions and Legionella growing on this [40]. Further research is required to gain a fuller understanding of this phenomenon. B. vesicularis has been used as a potential bioremediator of polyaromatic hydrocarbons [41]. All reported incidences of infection credited to B. vesicularis are listed in Table 1-3.

Identification of Brevundimonas spp
Members of the Brevundimonas spp. are Gram negative with cells appearing as straight slim rods upon Gram staining. They are non-spore forming. They are aerobic with optimal growth temperatures of between 30-37 C. They are oxidase positive and give variable results for catalase (usually positive). B. diminuta colonies have a chalk white appearance on MacConkey agar, whereas B. vesicularis colonies have an orange colour given by an intracellular pigment. Both grow slowly on ordinary nutrient media [42]. Both B. vesicularis and B. diminuta can be identified via commercial biochemical identification kits or systems such as the API 20 NE system, the VITEK 2 system (bioMerieux) or the Phoenix-100 automated system (Becton Dickinson). MALDI-TOF identification is also being used for identification of Brevundimonas spp. in clinical situations [43,44]. Species specific Real Time PCR primers and Fluorescence in situ hybridization (FISH) probes have been designed for B. diminuta [45]. These can be seen in Table 4.

Underlying causes
The majority of infections with Brevundimonas (Table 1-3) were found to have an underlying condition or disease that allowed patients to succumb to Brevundimonas infection. Seven patients, who were suffering with various types of cancer, contracted Brevundimonas -related bacteraemia, Urinary Tract Infection (UTI) and Empyema [46]; a 56-year-old female with Lupus glomerulonephritis acquired a Brevundimonas -related leg ulcer [47] and an infant suffering from Pompe disease was diagnosed with Brevundimonas -related bacteraemia [48]. Other examples of patients infected with Brevundimonas having underlying conditions are shown in Table 1-3. Such examples demonstrate the role of Brevundimonas as an opportunistic pathogen in immunocompromised individuals. Many of these instances of infection were hospital acquired although a large number were community acquired, which is interesting as opportunistic pathogens such as Brevundimonas spp or R. pickettii are usually contracted in hospital settings [7].

Co-Infection
Reports of cases of co-infection with Brevundimonas spp and other bacteria were rare with only two instances    [49].

Pseudo-outbreaks
As can be seen in Table 5 to date only two pseudo-outbreaks have been reported with Brevundimonas spp. Pseudo-outbreaks can be problematic as they can result in superfluous treatments given to patients (e.g. unnecessary antibiotics or the removal of indwelling devices such as catheters) and can waste valuable time and resources in the clinical setting. The causes of pseudo-outbreaks may be due to a number of different factors such as contaminated water used in the bacterial testing procedures or contamination of materials used in laboratory testing. Kim et al [50] described how B. diminuta was the cause of a pseudo-outbreak in a general hospital ward in South Korea. Patients did not display symptoms associated with bacterial infection, even though the organism was detected. The source of the B. diminuta contamination was not discovered. Lee et al. [51]. described B. diminuta as the cause of pseudo-outbreak in a tertiary care centre in the USA. The contamination was traced to pre-prepared inoculant media (used in the testing procedures for bacterial detection).

Treatment
The treatment of Brevundimonas spp. infections is frequently difficult, as these bacteria can be resistant to many different antibiotics including b-lactams and fluoroquinolones [46,47]. There have been no controlled trials of antimicrobial therapy for Brevundimonas spp. infections in humans therefore therapy should be informed by the results of in vitro susceptibility testing on isolates. In the majority of cases listed in Table 1-3 cephalosporins, penicillins or aminoglycoside antibiotics were given to treat patients and these were mostly successful.
Little is known about resistance mechanisms in Brevundimonas spp. Resistance to the fluoroquinolone family of antibiotics has been detected in outbreaks due to mutations in the quinolone resistance-determining region (QRDR) of the host gyrA, gyrB and parC genes [46]. Bla VIM-2 and Bla VIM-13 , which mediate resistance to almost all b-lactams (except aztreonam), have been found in both environmental and clinical isolates of B. diminuta [47]. The presence of Bla VIM-2 is related to a Tn1721-class 1 integron which was discovered in all B. diminuta isolates, with the determinant located on a plasmid [47]. This integron also had an aac (6 0 )-Ib gene, which mediates resistance to aminoglycoside antibiotics. Tetracycline resistance genes have also been found in environmental isolates of B. diminuta [52].

Breakdown of cases of infection with Brevundimonas spp.
Literature searches presented in Table 1-3 illustrate 49 separate instances of infection relating to Brevundimonas spp. The majority of these instances were infection with B. vesicularis (thirty-five cases -71%). One outbreak had both B. vesicularis and B. diminuta and one case of infection with B. vancanneytii was reported. The rest of the cases were made up B. diminuta infections (twelve cases-24%). The major breakdown of condition were as follows: seventeen instances of bacteraemia (34%), five instances of septicaemia/sepsis (10%), three instances of pneumonia/ pleuritis (6%), two instances each endocarditis (4%), keratitis (4%), and urinary tract infection (4%). Serious infections with Brevundimonas spp include four instances of septicaemia (8%), two of endocarditis (4%), one of septic arthritis (2%) and one of meningitis (2%). Other conditions include instances of two cases of tonsillitis (2%), two of liver abscess (2%) and two of botryomycosis (2%). There have also been two reported instances (4%) of Brevundimonas spp infection that have caused two or more conditions: bacteraemia and reactive arthritis and bacteraemia, urinary tract infection and empyema. Four instances of death have been related to Brevundimonas spp infection, three of bacteraemia and one of septicaemia.

Conclusions
Brevundimonas spp. are not currently considered as major pathogens. However, this should be re-evaluated in light of our investigations where forty-nine examples of Brevundimonas spp. infections have been found in the literature. These species have characteristics, such as ability to pass through sterilising filters, which may allow them to cause potentially harmful infections and even death on occasion. Although it is of low virulence and not as big a risk as other non-fermenting Gram-negative bacteria such as Burkholderia etc., it should not be over looked as a possible cause of nosocomial infections and should be considered for inclusion in hospital screening and prevention programs. These programs should consider investigation of possible Brevundimonas spp outbreaks if these bacteria are clinically isolated in more than one patient.