Soil Biology & Biochemistry 36 (2004) 539–543

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Short communication

Soil texture and irrigation influence the transport and the development
of Pasteuria penetrans, a bacterial parasite of root-knot nematodes
K.R. Dabiré*, T. Mateille1
UR IRD 51, Laboratoire de Nématologie, IRD, BP 1386, Dakar, Senegal
Received 22 June 2001; received in revised form 22 October 2003; accepted 27 October 2003

Abstract
The transport of the spores of Pasteuria penetrans was studied in three contrasted textured soils (a sandy, a sandy-clay and a clay soils),
cultivated with tomato, inoculated with juveniles of Meloidogyne javanica and watered with 25 or 150 mm day21. One month after
inoculation of the nematodes, 53% of the spores inoculated were leached by water flow in the sandy soil but only 14% in the sandy-clay soil
and 0.1% in the clay soil. No nematodes survived in the clay soil, while the population was multiplied both in the sandy and in the sandy-clay
soils. But juveniles of M. javanica were more infected by P. penetrans in the sandy-clay soil than in the sandy soil. Comparing different
combinations of bare soils containing 1.1– 57% of clay showed that the best spore percolation and retention balance occurred in soils
amended with 10 – 30% clay. However, the spore recoveries decreased when the soil was enriched with more than 30% clay. The role of clay
particles on the extractability of spores and on their availability to attach to the nematode cuticle in the soil is discussed.
q 2003 Elsevier Ltd. All rights reserved.
Keywords: Clay; Meloidogyne javanica; Pasteuria penetrans; Percolation; Soil texture

1. Introduction predict the success of P. penetrans inocula in nematode

Pasteuria penetrans, an endospore-forming bacterium,
was investigated as a biological control agent against plant-
parasitic nematodes, especially the root-knot nematodes
Meloidogyne spp. (Chen and Dickson, 1998). Some studies
were performed on the population dynamics of both
P. penetrans and Meloidogyne spp. in particular on
density-dependence (Ciancio and Bourijate, 1995). Few
papers have reported the effects of some abiotic factors,
such as temperature or soil moisture (Serracin et al., 1997)
and host plant (Giannakou et al., 1999) on the biological
control of Meloidogyne spp. populations. Research has also
been conducted on the influence of texture and ionic
solution on the availability of the spores that attach to the
nematode cuticle (Mateille et al., 1995). However, knowl-
edge about the survival and activity of the spores of
P. penetrans added to soils is lacking and is essential to
* Corresponding author. Address: DABIRE Kounbobr Roch,
INERA/Station de Farako-Bâ, BP 208, Bobo-Dioulasso Burkina Faso.
Tel.: þ 226-97-21-05; fax: þ 226-97-01-59.
E-mail address: dabire_roch@hotmail.com (K.R. Dabiré).
1
CBGP, Campus International de Baillarguet, CS 30016, 34988
Montferrier-sur-Lez Cedex, France.
0038-0717/$ - see front matter q 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/j.soilbio.2003.10.018
biological control management. The aim of this work is to
investigate the influence of (i) the soil texture on the
transport and the development of P. penetrans in cultivated
soils (ii) and the clay content on the transport and the
retention of spores in the bare soil.
2. Materials and methods
Three native soils were used (Table 1). A sandy soil
was sampled from a bare fallow at the experimental
station of the Institut Sénégalais de Recherches Agricoles
(ISRA, Cambérène, Sénégal). A clay soil was sampled
from a bare fallow at the west valley of the Senegal river.
Both soils were free of Meloidogyne spp. and of
P. penetrans. A sandy-clay soil was sampled from the
experimental station of the Ecole Nationale Supérieure
d’Agriculture (ENSA, Thiès, Senegal), in a field culti-
vated with African egg-plants (Solanum aethiopicum
Soxna), and highly infested with Meloidogyne javanica
(20 £ 103 juveniles cm21 of soil) and P. penetrans (80%
of infected juveniles). The soils were autoclaved (24 h at
120 8C) before using. Furthermore, different soil combinations
540 K.R. Dabiré, T. Mateille / Soil Biology & Biochemistry 36 (2004) 539–543

Table 1 the abundance of P. penetrans spores produced by

Physico-chemical characteristics of the soils used the nematode females, roots were dried, weighed and
Characteristics Sandy soil Sandy-clay soil Clay soil powdered. The root powders were suspended in 20 ml of
distilled water and the suspensions were sieved (20 mm).
Particles (%) The spores were numbered in the filtrates following the
Clay (0–2 mm) 1.1 10.3 57.0 technique for leachates as described above.
Fine silt (2– 20 mm) 0.9 1.7 12.4 A 25 g soil subsample was dispersed in 100 ml of
Coarse silt (20–50 mm) 1.2 2.4 10.0
distilled water in the presence of three agate marbles (1 cm
Fine sand (50–200 mm) 46.2 44 21.7
Coarse sand (200– 2000 mm) 50.4 40.7 0.2 diameter) for 16 h (Dabiré et al., 2001). After a 5 min
Organic matter (‰) sedimentation, the suspensions were sieved using a bank of
Carbon 0.24 0.29 2.68 sieves (200, 50, 20 mm). The size fraction 0 – 20 mm was
Nitrogen 0.04 0.04 0.42 concentrated on a 0.6 mm filter and suspended in distilled
C/N 6.00 7.25 6.38
water. The spores of P. penetrans were numbered as
Mineral analysis
P2O5 total (ppm) 198 1320 1477 described above and density was expressed per gram of soil
Ca (meq/%) 0.83 0.30 0.29 or per gram of root.
Mg (meq/%) 0.44 0.12 0.36 The juveniles of M. javanica were extracted from the
Na (meq/%) 0.17 0.14 0.22 remaining roots and soils (Seinhorst, 1962) and numbers
K (meq/%) 0.14 0.14 0.16
were estimated per plant. Multiplication rates of the
pH H2O 6.98 7.56 5.10
nematode populations were calculated as ratio of final
population (in roots and soil) to inoculum number. Five
replication were made per treatment.
were made with the native sandy and clay soils to obtain a
For the fallow soils, PVC tubes (10 cm high and
range of clay contents (Table 2).
1.5 cm internal diameter), closed at the bottom by a 50-
For the cultivated soils, PVC tubes (20 cm high and
mm mesh, were filled up to 9 cm height (Mateille et al.,
5 cm internal diameter), closed at the bottom by a 50-mm
1996) with 25 g of each native or mixed soils. The soils
mesh, were filled with each native soil (sand, sandy-clay
were saturated by submerging in distilled water for 20 min
and clay). The tubes were immersed in distilled water
and then allowed to drain. Then, 4.5 £ 106 spores of P.
during 20 min. After draining, 2-week-old tomato plants
penetrans were inoculated in the 0 – 1 cm top of the soil
(Lycopersicon esculentum Roma) were transplanted in
column. The tubes were placed on the top of 500 ml
each tube and randomly placed in a greenhouse (28 –
bottles under a drip distilled water supply. Different
30 8C). Each plant was inoculated with an average of
proportions of clay content were used (Table 2). After
8.5 £ 105 spores of P. penetrans two days later and with
24 h of percolation, the spores were numbered in the
about 180 juveniles of M. javanica another two days
leachates as described above. The soil columns were
later. Two irrigation treatments (25 or 150 ml of distilled
removed from the tubes and the soil was gently shaken by
water day21) were applied to the plants. In order to
hand in 150 ml of distilled water. After a 3– 5 min
recover the leachates, each tube was placed into a 150 ml
sedimentation, the soil suspensions were sieved using a
container. The leachates were sieved daily on a 0.6 mm
bank of sieves (200, 50, 20 mm). The spores were
filter and the spores of P. penetrans were numbered in
numbered in the size fraction 0 – 20 mm as described
the suspensions with a Malassez counting chamber
above.
(magnification 120).
Five replicates were used for each native or mixed soil
The plants were uprooted 30 days after nematode
and irrigation combination. Data were analysed according to
inoculation, and root galls induced by M. javanica were
ANOVA test. Percentages were transformed by Arcsin
indexed from 0 to 10 (Zeck, 1971). In order to evaluate
(sqrt) before analysis.

Table 2
Proportions of clay in the different soil combinations and irrigation
treatments
3. Results

Soil combinations (%) Clay particles (%) Irrigation (ml min-1) In the sandy soil, the irrigation did not influence the
root galling, the number of M. javanica juveniles in the
Sandy soil Clay soil
roots and the nematode multiplication rate (Table 3). But
100 0 1.1 100 the juveniles recovered from the soil were three time more
90 10 6.7 100 abundant under the 150 mm day21 irrigation ðP , 0:05Þ:
80 20 12.3 100 In the sandy-clay soil, the irrigation influenced the root
50 50 29.1 100 infestation and the multiplication rate, which were two
25 75 43.1 80
times highest under the 25 mm day21 irrigation (P , 0:05
0 100 57.0 80
for root infestation). In the clay soil, the inoculated
 K.R. Dabiré, T. Mateille / Soil Biology & Biochemistry 36 (2004) 539–543 541

Table 3
Influence of the soil type and the irrigation on the gall index, the soil and root infestations by M. javanica and the estimated reproduction of the nematode
population

Soil Irrigation (mm/day) Gall index Number of juveniles per plant Multiplication rate

In soil In roots

Sand 25 4a 1080a* 56,027a 334.1a

150 4a 3577b*** 50,738a 305.14a
Sandy-clay 25 4a 642c 83,100a 470.72a
150 4a 447c 45,720a 259.91a
Clay 25 1b* 3d 3b* 0.02b*
150 1b* 1d 0b* 0.01b*

Data within a column with the same letter are not significantly different, P , 0:05: Multiplication rate is final /inoculum number (178 juveniles).
*P , 0:05; **P , 0:01; ***P , 0:001:

juveniles did not survive whatever the irrigation applied.
Comparing the sandy and the sandy-clay soils, the former
had significantly high M. javanica densities but the roots
were similarly infested. The multiplication rates did not
differ under the 25 mm day21 irrigation. However, under
the 150 mm day21 irrigation, the multiplication rate
estimated for the sandy-clay soil was significantly lower
than for the sandy soil.
No spores were recovered in the percolates with
25 mm day21 watering whatever the soil. The spores
recovered in the leachate (Table 4) were four times more
abundant in the sandy soil than in the sandy-clay soil. Only a
few spores (about 40,000 spores) were recovered in the
leachate from the clay soil with 150 mm day21.
Irrespective of irrigation, more spores were recovered
from the clay soil than from both the sandy-clay and sand
soils.
Only a few spores were detected in the root powders
from the plants cultivated in the clay soil. The root
production of spores was highest in the plants cultivated in
the sandy-clay soil with 6 – 9 £ 105 spores g21 root
ðP , 0:025Þ: M. javanica juveniles were scarce in the
clay soil irrigated with 25 mm day21, but all of them were
infected by P. penetrans. In clay soils irrigated with
150 mm day21, they were absent. In the sandy soil, only
10% of juveniles were encumbered with spores, irrespec-
tive of the irrigation. The greatest infection of the juveniles
was observed in the sandy-clay soil, but the increased
irrigation reduced that significantly ðP , 0:05Þ:
After a 24 h percolation through the soil columns with
different clay contents, 76% of the inoculated spores
leached through the native sandy soil containing only
1.1% of clay particles (Fig. 1). The enrichment of the sandy
soil with the clay soil reduced the proportion of spores
detected in the percolate and no spores were leached
through the soils with more than 29% of clay. According to
the spore recovery from the soils, the optimal extraction
level (40% of the inoculated spores) was performed in soils
containing approximately 10– 12% of clay particles (includ-
ing the native sandy-clay soil). Fewer than 20% ðP , 0:05Þ
of inoculated spores were recovered from native clay soils
and from the soil mixtures containing more than 12% clay
particles.
4. Discussion
In sandy soils intensively irrigated, the loss of the
spores was ascribed to the continuous water pathway
which leached bacteria in depth. The phenomena were

Table 4
Influence of the soil and the irrigation on the leachate, the soil recovery and the root production of the spores of P. penetrans and on the proportion of
M.javanica infected juveniles

Soil Irrigation (mm/day) Spores of Pasteuria penetrans Infected juveniles in soil (%)
21 21
In leachate In soil (g ) In roots (g )

Sandy 25 0a 39 200a 612 676a 10.36a

150 2,275,000b*** 12,200a** 366,742a 10.24a
Sandy-clay 25 0a 207,200b 858,696a 64,73c***
150 630,188b* 88,400a 595,500a 27,51b**
Clay 25 0a 400,000b* 75,000b* 100c***
150 40,508a 228,000b 0b* 0a

Data within a column with the same letter are not significantly different, P , 0:05: *P , 0:05; **P , 0:01; ***P , 0:001:
542 K.R. Dabiré, T. Mateille / Soil Biology & Biochemistry 36 (2004) 539–543
Fig. 1. Proportions of spores of P. penetrans percolated and extracted in different native soils and soil mixtures defined by their clay content (bars represent
standard errors, *P , 0.05, **P , 0.025).
well elucidated by Wollum and Cassel (1978) studying
the loss of bacteria by percolation. Conversely, inten-
sively watered clay soils reduced the vertical movement
of spores. In our experiments this reduction occurred
when the sandy soils were supplemented with more than
29% of clay.
In these experiments, the retention of spores could be
attributed to the reduction of soil porosity consecutively
to the supplementation of clay particles. But, the
retention of bacteria can be secondarily due to adsorp-
tion on colloids when the soil structure is improved
abundantly with clay and organic matter (Bitton et al.,
1974). As it was previously noticed by Dabiré et al.
(2001), these adsorption phenomena are confirmed in our
experiments by the difficulty either to gently extract
spores of P. penetrans or to detect them in aqueous soil
suspensions from soils containing more than 12% of clay
particles. Therefore, according to our results, two clay-
content thresholds can be defined with regard to the best
balance between percolation (about 10% of clay) and
retention (about 30% of clay) of the spores in the soils.
These soil interactions could directly control the
dispersal and the availability of the spores of P.
penetrans to adhere on nematode cuticle. Therefore, it
is predicted that biocontrol of plant parasitic nematodes
could be most effective in soils with 10 –30% of clay
particles.
The increase of the M. javanica populations was
enhanced in both sandy and sandy-clay soils, confirming
that the sandy and sandy-clay soils offer more suitable
abiotic conditions, such as aeration, which increased
juvenile hatching (Van Gundy, 1985). However, water
saturation induces a nematode population decline
by oxygen depletion especially in the clayey soils
(Prot and Van Gundy, 1981) which accords with our
results obtained in clay soils.
The attachment of P. penetrans spores to the
nematode cuticle is humidity dependent. In sandy soils
characterized by low water holding capacity (Bonneau
and Souchier, 1979), a 25 ml irrigation did not provide
enough moisture for spore attachment. Furthermore,
when this soil was adequately watered, the populations
of Meloidogyne increase while the bacteria were mostly
leached down, resulting in a dilution of the infective
spores. In comparison, the sandy-clay soil provides more
juvenile infection, revealing that its abiotic properties
(water holding capacity, aeration) are more suitable for
juvenile infection and P. penetrans development.
Acknowledgements
This study was supported by a grant from the EC
Project STD 3 no. TS3 * CT92-0098 (Biocontrol of
damaging root-knot nematode (Meloidogyne spp.) pests of
staple food and cash crops by including suppressive soils
with the bacterial parasite P. penetrans). Authors are
grateful to anonymous reviewers for their helpful sugges-
tions and comments.
References
Bitton, G.N., Lahav, G.N., Henis, Y., 1974. Movement and retention of
Klebsiella aerogenes in soil columns. Plant Soil 40, 373–380.
Bonneau, M., Souchier, B., 1979. Pédologie 2: Constituants et propriétés du
sol (Eds.). Masson Paris, 459 pp.
 K.R. Dabiré, T. Mateille / Soil Biology & Biochemistry 36 (2004) 539–543
Chen, Z.X., Dickson, D.W., 1998. Review of Pasteuria penetrans: biology,
ecology and biological control potential. Journal of Nematology 30,
313–340.
Ciancio, A., Bourijate, M., 1995. Relationship between Pasteuria penetrans
infection levels and density of Meloidogyne javanica. Nematology
Mediteranean 23, 43 –49.
Dabiré, K.R., Chotte, J.L., Fardoux, J., Mateille, T., 2001. New
development in the estimation of spores of Pasteuria penetrans.
Biology and Fertility of Soils 33, 340– 343.
Giannakou, I.O., Pembroke, S.R., Gowen, S.R., Douloumpaka, S., 1999.
Effects of fluctuating temperatures and different host plants on
development of Pasteuria penetrans in Meloidogyne javanica. Journal
of Nematology 31, 312–318.
Mateille, T., Duponnois, R., Diop, M.T., 1995. Influence des facteurs
telluriques abiotiques et de la plante hôte sur l’infection des nématodes
phytoparasites du genre Meloidogyne par l’actinomycète parasitoı̈de
Pasteuria penetrans. Agronomie 15, 581–591.
Mateille, T., Duponnois, R., Dabiré, K., Diop, M.T., Ndiaye, S., 1996.
Influence of the soil on the transport of the spores Pasteuria penetrans,
543
parasite of the genus Meloidogyne. European Journal of Soil Biology
32, 81–88.
Prot, J.C., Van Gundy, S.D., 1981. Effect of soil texture and the clay
component on migration of Meloidogyne incognita juveniles. Journal of
Nematology 13, 213–217.
Serracin, M., Schuerger, A.C., Dickson, D.W., Weingartner, D.P., 1997.
Temperature-dependent development of Pasteuria penetrans in Meloi-
dogyne arenaria. Journal of Nematology 29, 228– 238.
Seinhorst, J.W., 1962. Modifications of the elutriation method for
extracting nematodes from soil. Nematologica 8, 117–128.
Van Gundy, S.D., 1985. Ecology of Meloidogyne spp: emphasis on
environment factors affecting survival and pathogenicity. In: Barker,
K.R., Carter, C.C., Sasser, J.N. (Eds.), An Advanced Treatise on
Meloidogyne. Vol. I. Biology and Control, IMP, North Carolina State
University Graphics, USA, pp. 177–182.
Wollum, A.G., Cassel, D.K., 1978. Transport of microorganisms in sand
columns. Soil Science Society American Journal 42, 72– 76.
Zeck, W.M., 1971. A rating scheme for field evaluation of root-knot
nematode infestations. Pflanzen-Nachricht. Bayer Ag. 24, 141– 144.