Variability of Burkholderia pseudomallei Strain Sensitivities to Chlorine Disinfection

Abstract

Burkholderia pseudomallei is a select agent and the causative agent of melioidosis. Variations in previously reported chlorine and monochloramine concentration time (Ct) values for disinfection of this organism make decisions regarding the appropriate levels of chlorine in water treatment systems difficult. This study identified the variation in Ct values for 2-, 3-, and 4-log 10 reductions of eight environmental and clinical isolates of B. pseudomallei in phosphate-buffered water. The greatest calculated Ct values for a 4-log 10 inactivation were 7.8 mg ⅐ min/liter for free available chlorine (FAC) at pH 8 and 5°C and 550 mg ⅐ min/liter for monochloramine at pH 8 and 5°C. Ionic strength of test solutions, culture hold times in water, and cell washing were ruled out as sources of the differences in prior observations. Tolerance to FAC was correlated with the relative amount of extracellular material produced by each isolate. Solid-phase cytometry analysis using an esterase-cleaved fluorochrome assay detected a 2-log 10 -higher level of organisms based upon metabolic activity than did culture, which in some cases increased Ct values by fivefold. Despite strain-to-strain variations in Ct values of 17-fold for FAC and 2.5-fold for monochloramine, standard FAC disinfection practices utilized in the United States should disinfect planktonic populations of these B. pseudomallei strains by 4 orders of magnitude in less than 10 min at the tested temperatures and pH levels.Burkholderia pseudomallei, a gram-negative soil saprophyte, is endemic to Southeast Asia (30) and northern Australia and has been linked to waterborne illness in these areas (9, 15). B. pseudomallei has been identified by the Centers for Disease Control and Prevention (CDC) as a select agent. The organism can tolerate a wide variety of soil and temperature conditions (33), which enables it to exist in soil and water in subtropical zones (10,19,32,35). Information on survival of B. pseudomallei in drinking water containing disinfectants is necessary to ensure that the end users of water distribution systems are sufficiently protected from exposure to this organism.Free available chlorine (FAC) and monochloramine are the most commonly used drinking water disinfectants (1, 2, 28). Factors such as ionic strength, pH, turbidity and chlorine demand, biofilm growth, and the physiological state of the cells can alter disinfection efficacy (6,16,18,22,31). The contact concentration and time values (Ct) (mg ⅐ min/liter) required to achieve disinfection have been observed to vary between strains of the same organism (23, 36). Additionally, stressrelated phenotypic changes in Vibrio species have been linked to increased tolerance to chlorine (20,24,34).Several strains of B. pseudomallei endemic to northern Australia were found by Howard and Inglis (13) to be more chlorine and monochloramine tolerant than other strains via techniques such as plate culture, a most-probable-number (MPN) recovery method, and flow cytometry using a membrane integrity stain. Disinfectio...

Inactivation of Brucella Suis, Burkholderia pseudomallei, Francisella tularensis, and Yersinia pestis using Vaporous Hydrogen Peroxide

“…Previously, both F. tularensis and Y. pestis dried onto non-porous surfaces were completely inactivated by vaporous hydrogen peroxide in chambers ranging in volume up to approximately 3,600 liters . Brucella suis and B. pseudomallei have been shown to be inactivated by freechlorine, monochloramine and ultraviolet light (O'Connell et al, 2009;Rose et al, 2007;Sagripanti et al, 2009); however, this study provides the first demonstration of inactivation by using vaporous hydrogen peroxide, especially on a large-scale.…”

Using Amplicon Sequencing To Characterize and Monitor Bacterial Diversity in Drinking Water Distribution Systems

“…For example, the Burkholderiales, which are known to contain strains resistant to antibiotics (27), were identified in high percentages in the postdisinfection samples in both systems. The sensitivity of Burkholderia to chlorine (28) and monochloramine (29) is known to vary considerably, with some isolates more tolerant than others. Previous research has revealed that cocultures of the pathogenic Burkholderia species Burkholderia pseudomallei with the ubiquitous freshwater amoeba Acanthamoeba astronyxis greatly enhances the survival of B. pseudomallei in the presence of monochloramine, with 100 times more monochloramine required to maintain disinfectant efficacy when the amoeba was present (29).…”

Inactivation of bacterial biothreat agents in water, a review

“…There appears to be much variation in tolerance to disinfectants within this species, though little is known about the resistance mechanism (Howard & Inglis 2003, 2005; O’Connell et al 2009). Some strains produce increased amounts of mucoid polysaccharide, which is readily observed in their colonial morphology, and has been reported to affect resistance to UV light, but was not directly correlated to FAC resistance (Howard & Inglis 2005).…”

“…One study conducted with Australian isolates found some cells in a test suspension survived 1,000 mg L −1 FAC, using a broth-based most probable number (MPN) culture method (Howard & Inglis 2003). In contrast, using the same MPN culture method as well as a standard plate count culture method, O’Connell et al (2009) tested 11 strains of various origins and morphologies (but not the same Australian isolates mentioned previously) and found that all strains were inactivated within 10 minutes with a FAC concentration of 1 mg L −1 (Table 2). These findings suggest that a wide range of susceptibility exists within the species.…”