1. Burkholderia pseudomallei in a rice paddy in Northern Laos - influence of depth, season and physicochemical parameters
Poster PPV-03
MANIVANH L.1, PIERRET A.2, RATTANAVONG S.3, BUISSON Y.1, ELLIOTT I.3, MAEGHT J.-L.2, XAYYATHIP K.2, VONGSOUVATH M. 3, PHETSOUVANH R.3,4, NEWTON P.N.3.4, DANCE D.A.B.3,4
1 - Institut de la Francophonie pour la Médecine Tropicale, BP 9519, Vientiane, Lao People’s Democratic Republic
2 - Institut de Recherche pour le Développement, Bioemco, BP 5992, Vientiane, Lao People’s Democratic Republic
3 - Lao-Oxford-Mahosot Hospital-Wellcome Trust Research Unit, Microbiology Laboratory, Mahosot Hospital, Vientiane, Lao People’s Democratic Republic
4 - Centre for Clinical Vaccinology and Tropical Medicine, Churchill Hospital, University of Oxford, Oxford, United Kingdom
Figure 1
Background and aim
The presence of Burkholderia pseudomallei in Lao soil was first demonstrated in
Human melioidosis has been recognised in Laos since
The true incidence and distribution across the country are unclear due to a lack of diagnostic facilities.
Although B. pseudomallei is known to be focally distributed in the environment in some tropical regions, the factors underlying its distribution are poorly understood.3
To gain insight into the influence of physico-chemical factors on its distribution in Laos, we have intensively studied a single rice paddy over a 12-month period. 1 1 1
Materials and methods
Study site, sampling strategy and soil analyses
Study site in Nabon village, Phonhong district (Vientiane province), identified as contaminated with B. pseudomallei in 1998 (Figure 1)1.
Soil samples collected on 4 occasions (end of dry season; onset of rainy season; end of rainy; end of second dry season)
Samples collected from surface and depths of 30, 60 and 90cm (when possible) from each of 49 5 x 5 m squares.
Samples cultured semi-quantitatively for B. pseudomallei by procedure described by Rattanavong et al.3 and Vongphayloth et al4.
Soil parameters measured include:1) soil bulk density, 2) water content, 3) particle size , 4) pH, 5) temperature, 6) soil organic matter (SOM), 7) total C and N by dry combustion, 8) extractable P (Olsen method), 9) Cu, Zn, Fe, Mn (EDTA extractable), 10) Ca, Mg, Mn, Al, Na, K (cobaltihexammine), 11) cation exchange capacity (CEC, Metson), and 12) black carbon.
Figure 2
Results
At the end of 2011 dry season 33% of samples contained B. pseudomallei. After first monsoon rains, only 19% of samples were positive. By the end of 2011 rainy and 2012 dry seasons, 28% and 32% of samples, respectively, contained B. pseudomallei (Fig. 2).
In dry seasons, there was a positive gradient of B. pseudomallei occurrence with depth, from <10% near soil surface to >50% at 90 cm (p <0.001). This reversed in the rainy season, with more positive samples at 5 and 30 cm and fewer at 60 cm. Bacterial densities in positive samples also increased during the rainy season.
Below 30 cm, non-significant positive associations with CEC, SOM and pH were observed; soil parameters were unrelated to the presence/absence of B. pseudomallei.
Aggregating data from the four sampling campaigns shows that, irrespective of soil depth, positive samples at >100 CFU/g were located in wetter soil than negative samples and samples positive at <100 CFU/g (Welsh two sample t-test, p<0.05) (Fig. 3).
At 90 cm, >50% of the positive samples occurred within the same 5 x 5 m sampling square at the end of the two dry seasons (Fig. 4).
Figure 3
Figure 4
Conclusions
Similar bacterial distributions at the end of the two dry seasons, at 90 cm; soil water content and numbers of B. pseudomallei positively associated
⇒ Existence of deep, specific soil micro-sites (reservoirs), from which B. pseudomallei rises to the surface with the rains?
But… no statistically significant associations between soil physicochemical parameters and numbers of B. pseudomallei.
Sensitivity of enrichment culture (broth containing crystal violet and colistin (50 mg/l)) only 43% of that of direct culture of diluted soil suspension on Ashdown’s agar,
⇒ overgrowth by competing flora?
⇒ Further studies needed to optimise detection methods for B. pseudomallei in Lao soil.
1 Wuthiekanun et al. (2005). J Clin Microbiol 43:
2 Phetsouvanh et al. (2001). Clin Infect Dis 2001; 32:
3Cheng & Currie (2005) Clin Microbiol Rev 18:
4Ratanavong et al. (2011). Appl Environ Microbiol 77:
5Vongpayloth et al. Am J Trop Med Hygiene (in press)