METABOLISM AND NUTRITION

Mortality and Growth Performance of Broilers Given Drinking Water

Supplemented with Chicken-Specific Probiotics

H. M. Timmerman,*† A. Veldman,‡ E. van den Elsen,§ F. M. Rombouts,# and A. C. Beynen*1

*Department of Nutrition, Faculty of Veterinary Medicine, Utrecht University, PO Box 80152, 3508 TD Utrecht,
The Netherlands; †Research and Development Department, Winclove Bio Industries B.V., PO Box 37239,
1030 AE Amsterdam, The Netherlands; ‡Schothorst Feed Research, PO Box 8200 AM Lelystad, The Netherlands;
§Research and Development Department, Fransen mengvoeders B.V., PO Box 30, 5469 ZG Erp, The Netherlands; and
#Laboratory of Food Microbiology, Department of Agrotechnology and Food Sciences,
Wageningen University, PO Box 8129, 6700 EV Wageningen, The Netherlands

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ABSTRACT For application in broiler production, we
developed a multispecies (MSPB) and a chicken-specific
(CSPB) probiotic preparation in fluid form. The MSPB
contained different probiotic species of human origin,
whereas the CSPB consisted of 7 Lactobacillus species iso-
lated from the digestive tract of chickens. In a field trial
with broilers, MSPB treatment resulted in a slight increase
(by 1.84%) in broiler productivity based on an index tak-
ing into account daily weight gain, feed efficiency, and
Key words: broiler, multispecies probiotic, chicken-specific probiotic, feed efficiency
mortality. The CSPB treatment reduced mortality in 2
subsequent field trials and raised productivity by 2.94
and 8.70%. In a controlled trial with broilers showing a
high index of productivity, probiotic treatment further
raised productivity by 3.72%. Based on the present 4 stud-
ies in combination with 9 studies published earlier, it is
suggested that with higher productivity rates of the broil-
ers the effect of probiotics becomes smaller.
2006 Poultry Science 85:1383–1388

INTRODUCTION
Colonization of the digestive tract with commensals
induces in the host a set of genes associated with postnatal
maturation, mucosal barrier fortification, innate immune
responses, and promotion of nutrient metabolism
(Hooper et al., 2001; Stappenbeck et al., 2002; Rawls et
al., 2004). A well-accepted method to quickly introduce
a commensal microflora in hen-deprived chicks is
through the administration of probiotics. The most widely
used probiotic strains are of the genus Lactobacillus, which
is also the dominant genus of the proximal intestine of
chickens early in life (Barnes et al., 1972). Edens et al.
(1997) showed that in ovo and ex ovo administration of
Lactobacillus reuteri resulted in an increased villus height,
indicating that probiotics are potentially able to enhance
nutrient absorption and thereby improve growth perfor-
mance and feed efficiency. Indeed, Lactobacillus adminis-
tration has been shown to improve growth rates and feed
(1997) showed that in ovo and ex ovo administration of
(Jin et al., 1998b, 2000; Zulkifli et al., 2000; Kalavathy et
al., 2003). Through so-called competitive exclusion, probi-
otics are potentially able to reduce mortality from enteric
pathogens, but in trials with broilers given Lactobacillus
preparations the effects on mortality were inconsistent
(Watkins and Kratzer, 1984; Jin et al., 1998a,b, 2000; Zul-
kifli et al., 2000). However, in challenge experiments with
pathogens such as Escherichia coli, Salmonella typhimurium,
and Staphylococcus aureus, there was a probiotic-induced
reduction in mortality (Watkins and Miller, 1983; Edens
et al., 1997).
We have provided evidence that multispecies probiot-
ics (MSPB) are more effective than monospecies probiot-
ics (Timmerman et al., 2004) and also that species-specific
probiotics elicit different health effects than do probiotics
derived from another host species (Timmerman et al.,
2005). To further qualify the potential of probiotics to
improve growth performance and mortality in broilers,
we investigated the effect of a chicken-specific probiotic
(CSPB) that was administered with the drinking water.
To evaluate the application of the CSPB in practice, the
efficacy was not only studied in a controlled experiment
but also in 2 field trials. In an additional field trial we
used a MSPB containing different probiotic species of
human origin. To perform the 4 experiments, we devel-
oped a fermentation medium that is suitable for adminis-
tration via the drinking water, thereby rendering redun-
dant the use of expensive freeze-dried preparations.
MATERIALS AND METHODS

Probiotic Strains
2006 Poultry Science Association, Inc.
Received July 20, 2005.
Accepted December 8, 2005. For this study 2 different liquid probiotic formulas were
1
Corresponding author: a.c.beynen@vet.uu.nl developed. The MSPB preparation contained commer-

1383
1384 TIMMERMAN ET AL.
cially available probiotic strains. A combination of 6
strains was used: Lactobacillus acidophilus W55, Lactobacil-
lus salivarius W57, Lactobacillus casei W56, Lactobacillus
plantarum W59, Lactococcus lactis W58, and Enterococcus
faecium W54 (Winclove Bio Industries B.V., Amsterdam,
The Netherlands). To formulate the CSPB preparation,
Lactobacillus strains were isolated from fresh digesta and
intestinal tissue samples taken from healthy chickens and
were screened for probiotic properties.
Isolation of Lactobacillus Strains from Chickens.
Digesta and tissue samples of the crop, small intestine,
and cecum were collected freshly from layer hens and
broiler chickens and were stored in sterile, buffered pep-
tone water (Oxoid, Haarlem, The Netherlands). The sam-
ples were kept refrigerated and were processed on the
same day. The digesta samples were homogenized in
buffered peptone water with an Ultra Turrax blender
(Janke and Kunkel, IKA Labortechnik, Staufen, Germany)
under anaerobic conditions. Mucosal scrapings were de-
rived from the tissue samples and dissolved in buffered
peptone water. Further processing was performed under
aerobic conditions. Serial dilutions of the homogenized
samples were made in reduced physiological salt solution
made as follows (grams per liter): neutralized bacteriolog-
ical peptone, 1; l-cysteine-HCl, 0.5 (Sigma-Aldrich
Chemie GmbH, Steinheim, Germany) and NaCl, 8 (Acros
Organics, Geel, Belgium). Lactobacillus strains were cul-
tured on LAMVAB plates selective for Lactobacilli due to
low pH (5.0) and the presence of vancomycin in the me-
dium (Hartemink et al., 1997). The plates were incubated
anaerobically (Anoxomat, Mart, Lichtenvoorde, The
Netherlands) at 37°C for 48 h.
Well-isolated colonies with different appearances were
picked from each plate and transferred to new LAMVAB
plates and further incubated at 37°C for 48 h (anaerobic)
to obtain pure strains. Finally, 150 pure colonies were
isolated and grown in de Man, Rogosa, and Sharpe broth
(Merck, Darmstadt, Germany) at 37°C for 24 h. Aliquots
of these cultures were stored with 30% glycerol at −80°C.
All strains were screened for useful properties to produce
a liquid probiotic supplement, as described before (Tim-
merman et al., 2005). In short, strains were tested for
growth rate at different pH, acidification rate, and inhibi-
tion of pathogens like E. coli and S. typhimurium. Based
on these results 23 promising strains were selected for
identification by fermentation patterns with standard An-
alytical Profile Index (API 50CHL, BioMérieux, Inc., Ha-
zelwood, MO) tests. The selection of strains was then
checked for growth and stability, as assessed by viable
cell count after 2 wk of refrigerated storage, in a liquid
fermentation medium (see later in text). Based on these
results, the following 7 Lactobacillus strains were selected
for composing the CSPB: Lactobacillus bifermentans
W204.5, Lactobacillus sanfranciscensis W205.6, Lactobacillus
sanfranciscensis W208.6, Lactobacillus reuteri W218.2, Lacto-
bacillus reuteri W223.5, Lactobacillus reuteri W227.3, and
Lactobacillus fermentum W227.5. The selected isolates with
their identification and screening results are presented in
Table 1.
Formulation of the MSPB and CSPB Preparations.
The liquid growth medium was composed of 200 g of
soy protein hydrolysate (Heybroek, Amsterdam, The
Netherlands), 160 g of yeast extract (Gistex LS powder
AGGL, DSM Food Specialties, Delft, The Netherlands),
200 g of dextrose (Roquette, www.roquette.com), and 80
g of minerals [a combination of potassium chloride, mag-
nesium sulphate, and manganese sulphate (Kortex, The
Netherlands)]. These ingredients were dissolved in 15 L
of hot tap water, and 25 L of cold tap water was then
added. The product was cooled down quickly and stored
refrigerated in small containers. For formulation of both
the MSPB and CSPB preparation, the component strains
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(pregrown in de Man, Rogosa, and Sharp broth and stored
at −80°C) were first individually inoculated at 10 mL/L
and grown overnight at 37°C in the liquid fermentation
medium.
After fermentation, the pH of the cultures was deter-
mined (pH < 4.2) and the optical density of the cultures
was measured using a spectrophotometer (λ = 620 nm)
to assure growth of all strains (optical density > 1.00).
Also, samples were taken for viable cell count analysis
of each strain. Then, equal volumes of each of the 6 cul-
tures were mixed and a sample was taken to determine
the total viable cell count of the finished product (∼109
cfu/mL). The product was kept refrigerated and was
stored for up to 2 wk. During storage the total cell count
was checked regularly to verify the stability of the
product.
Because of administration problems of the probiotic
preparation in field trials 1 and 2 (see later), attempts were
made to optimize the probiotic preparation to prevent
clogging and sedimentation of probiotic components in
the drinking water system in field trial 3. Essentially, the
procedure of preparation remained unchanged, except
that instead of adding the whole fermentation broth to
the drinking water, the probiotic cells were first washed
with buffered peptone water by use of crossflow filtration
(Sartorius Technologies B.V., Nieuwegein, The Nether-
lands). During this procedure the final concentration of
the product was increased to 1.0 × 1010 cfu/mL.
Experimental Design
Field Trial 1. This trial took place at the research station
of Fransen Mengvoeders B.V. Five thousand 1-d-old male
Cobb broiler chicks (Cobroed, Lievelde, The Netherlands)
were housed in a standard broiler house divided in 2
similar areas with separate feed and drinking facilities.
Broilers were fed a commercial starter diet from 0 to 14
d of age (ME, 2,850 kcal/kg; CP, 220 g/kg), a grower diet
from 14 to 28 d (ME, 3,025 kcal/kg; CP, 202 g/kg), and
a finisher diet from d 29 onward (ME, 3,050 kcal/kg; CP,
192 g/kg). The anticoccidial antibiotics nicarbazin (125
mg/kg) and salinomycin (70 mg/kg) were added to the
starter and grower diets, respectively. Water and feed
were supplied for consumption ad libitum. The MSPB
treatment was randomly assigned to one-half of the house
and started immediately after arrival of the chicks. Two
 EFFECT OF PROBIOTICS ON MORTALITY AND GROWTH 1385
Table 1. Selected Lactobacillus species for the preparation of the liquid chicken-specific probiotic preparation

Pathogen inhibition3
Growth rate at different pH levels1
Listeria Escherichia Salmonella
Isolate pH 5.6 pH 4.0 Acidification2 monocytogenes coli typhimurium Identification4

W204.5 ++ ++ + 15.5 15.0 17.3 Lactobacillus bifermentans

W205.6 ++ ++ ++ 17.3 17.0 17.5 Lactobacillus sanfranciscensis
W208.6 ++ ++ + 15.3 17.0 15.3 Lactobacillus sanfranciscensis
W218.2 ++ ++ ++ 14.5 15.5 13.8 Lactobacillus reuteri
W223.5 ++ ++ ++ 15.5 14.3 13.5 Lactobacillus reuteri
W227.3 ++ + ++ 23.0 17.0 20.8 Lactobacillus reuteri
W227.5 ++ + ++ 23.5 17.0 18.5 Lactobacillus fermentum
1
Growth rate at different pH levels was assessed as short lag phase, followed by rapid growth in the exponential phase (steep slope) and a high
total cell count at the end of the experiment (measured by optical density), designated as (++). Strains with (+) showed a lower optical density
value at the end of the experiment. The experiment was performed 3 times for each strain.

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2
Acidification was assessed as a change in color (at γ = 405 nm in the microplate reader) during 32 h of the pH-indicators bromo-cresol green
and purple. A fast and high increase is indicated with ++, a slower or less intensive increase is indicated with +. The experiment was performed
3 times for each strain.
3
Inhibition is measured as the diameter (mm) of the inhibition zone for all pathogens. The values in the table are the average of 2 measurements.
4
Identification according to the carbohydrate fermentation profile (API 50 CH, BioMérieux, Inc., Hazelwood, Mo).

thousand five hundred broiler chicks were administered
the MSPB via the drinking water from d 0 to 14 at a dose
rate of 10 mL per liter of drinking water by use of a
dosage pump (Aquados, VLM B.V., Mariahout, The Neth-
erlands). The total amount of MPSB administered was
adjusted daily based on an expected growth curve to
achieve an average dose of 2.0 × 109 cfu/kg of BW.
Field Trial 2. After carrying out the first field trial
the CSPB preparation was developed. The design of the
experiment was similar to that of field trial 1 except for
the following modifications. Immediately after arrival,
the chicks of the probiotic-treated group were sprayed
with a diluted CSPB preparation, which resulted in ap-
proximately 4 × 107 probiotic organisms per sprayed
chick. Two thousand six hundred ten out of the 5,220 1-
d-old broiler chicks were administered the CPSB via the
drinking water from d 0 to 31 at an approximate rate of
4 × 108 cfu/kg of BW.
Field Trial 3. This experiment was conducted as field
trial 2 except that the CSPB preparation was concentrated
by means of crossflow filtration as just described. Dosage
and duration of treatment were as in field trial 1.
Controlled Trial. The controlled trial took place at the
research station of Schothorst Feed Research. Four hun-
dred and twenty 1-d-old female broiler Ross 508 (Co-
broed, Lievelde, The Netherlands) chicks were randomly
divided into 12 groups of 35 chicks each. Each group was
assigned to a floor pen that contained a self-feeder and
waterer to provide ad libitum access to feed and water.
Broilers were fed a commercial starter diet from 0 to 14
d of age (ME, 2,775 kcal/kg; CP, 205 g/kg), a grower diet
from 14 to 30 d, and a similar finisher diet from 31 to 37
d (ME, 2,900 kcal/kg; CP, 200 g/kg). The anticoccidial
antibiotics diclazuril (1 mg/kg) and monensin (100 mg/
kg) were added to the starter and grower diets, respec-
tively. Probiotic treatment was randomly assigned to 6
groups of 35 chicks. Spraying and supplementation rates
of CSPB were identical to those described for field trial 2.
Data Collection
In field trials 1–3, BW was recorded only at the start
and at the end of the experiment. Body weight at d 0 was
assessed as the average of a random selection of 250 chicks
per experimental flock. Final BW was assessed by divid-
ing the total weight per experimental flock by the number
of chicks alive before transportation to the processing
plant. Feed and water consumption on a flock basis was
recorded daily. In the controlled trial, weight gain and
feed intake were recorded for 3 growth stages (starter: d
0 to 14; grower d 14 to 30; and finisher: d 30 to 39). In
all trials mortality was recorded daily, and percentage

Table 2. Growth performance and mortality of broilers in 3 pooled field trials and a controlled trial with administration of a multispecies probiotic
(MSPB) or a chicken-specific probiotic (CSPB) in the drinking water

Pooled data field trials1 Controlled trial2

Treatment Control Probiotic SEM P-value Control CSPB SEM P-value

Final BW, g 2,357 2,342 43.1 0.410 1,978 2,003 18.9 0.184
Feed intake, g 4,460 4,300 94.6 0.149 3,153 3,246 47.38 0.097
Average daily gain, g/day 49.99 49.65 0.61 0.358 52.4 53.1 0.51 0.184
Feed conversion, kg of feed/kg of gain 1.93 1.87 0.02 0.044 1.66 1.67 0.01 0.116
Flock total BW, kg 5,448 5,506 101.0 0.354 385.8 404.6 9.92 0.105
Mortality rate, % 8.84 7.27 0.59 0.066 7.14 3.81 1.91 0.197
1
Data are means of 3 field trials. In field trial 1 the MSPB was applied. In field trials 2 and 3 the CSPB was applied.
2
Data are means of 6 replicate pens of 35 birds each.
1386
Figure 1. Cumulative mortality of broilers fed either a multispecies probiotic (MSPB).
mortality was calculated. Feed conversion was calculated
per experimental unit as total feed intake (kg): total gain
of live chickens (kg).
Statistical Analysis
The data for each variable were subjected to 1-way
ANOVA (Steel and Torrie, 1980). Differences between
treatment groups were evaluated with the Student’s t-
test using the GLM procedure of SAS (SAS Institute, 2000).
The level of statistical significance was preset at P < 0.05
for 1-sided testing. Based on literature (Watkins and
Kratzer, 1984; Jin et al., 1998a,b, 2000; Abdulrahim et
al., 1999; Kalavathy et al., 2003) it was expected that the
treatment effect goes in 1 direction. Mortality within ex-
periments was evaluated by means of Fisher’s exact test.
RESULTS
Growth performance and mortality for all field trials
are pooled in Table 2. Figure 1 shows the time course of
cumulative mortality. In field trial 1, administration of
TIMMERMAN ET AL.
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the MSPB preparation had no effect on weight gain and
feed conversion (data not shown). Overall mortality was
similar for the control and MSPB-treated groups, but Fig-
ure 1 illustrates that until 42 d the mortality was lower in
the MSPB group. During the last week of the experiment
ambient temperature was extremely high, which proba-
bly caused the rapid increase in mortality in both groups.
The increase was greater in the broilers given MSPB so
that overall mortality in the 2 groups became similar. In
field trial 2 the newly developed CSPB was used. The
initial progress of the trial was complicated by an E. coli
infection of the yolk sac, which became evident on d 2.
There was a sharp rise in initial mortality. When com-
pared with the control birds, the probiotic-treated chicks,
which were kept in the same house, were visually less
affected by the infection. During the first 3 d, 27 chicks
in the probiotic-treated group died compared with 73
in the control group. To prevent excessive mortality, all
animals were treated with a combination of sulfamethox-
azole (80%) and trimethoprim [20%; T.S. SOL (Dopharma,
www.dopharma.com)] from d 3 to 6. During antibiotic
treatment, probiotic treatment was stopped because in-
 EFFECT OF PROBIOTICS ON MORTALITY AND GROWTH 1387
vitro experiments had shown that the CSPB strains were Table 3. Production numbers for broilers in the current studies and in
published studies as affected by probiotic treatment
highly sensitive to the antibiotic used. After antibiotic
treatment, probiotic treatment was reinstalled. The CSPB Production number1 change
treatment caused a slight decrease in overall mortality as caused by probiotic treatment
and also produced an improvement of feed conversion Description Control Absolute %
(data not shown). In field trials 1 and 2 it was observed Current study
frequently that insoluble fermentation metabolites sedi- Field trial 1 245.3 +4.53 +1.84
mented and subsequently clogged the dosage pump. In Field trial 2 242.6 +7.12 +2.94
Field trial 3 247.6 +21.5 +8.70
the controlled trial, the results of which are given later, Controlled trial 309.8 +11.5 +3.72
the chicks given drinking water with CSPB drank less. Literature2
Possibly, this was related to an adverse taste response of 1a 126.2 +11.4 +9.01
1b 126.2 +34.4 +25.0
the water due to the organic acids present in the CSPB 2a 214.3 +28.6 +13.3
preparation. Neutralization of the acids with calcium car-

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bonate raised water intake. It was decided to separate
the medium and the probiotic cells by washing the cells
with buffered peptone water as described in the Materials
and Methods section. The improved CSPB preparation
was used in field trial 3, and the technical problems seen
in the earlier experiment were prevented. Probiotic treat-
ment resulted in a clear improvement of the feed conver-
sion (data not shown) and a decrease in mortality.
The design of the field trials was such that in each trial
there was only 1 experimental unit per treatment. To
carry out statistical analysis, the data of the 3 field trials
were pooled even though different probiotic preparations
had been used. After pooling of the data, probiotic treat-
ment appeared to have caused a statistically significant
improvement of feed conversion. The probiotic-induced
decrease in overall mortality just failed to reach statisti-
cal significance.
A controlled trial was carried out with the probiotic
preparation used in field trial 2. The results are presented
in Table 2. The treatment with CSPB decreased feed con-
version in the starting period (d 0 to 14; data not shown).
Average daily gain during the growth phase (14 to 30 d)
and BW at 30 d were significantly higher in the CSPB-
treated group (data not shown). This growth-promoting
effect was not present at the end of the trial. The CSPB-
mediated reduction in mortality was similar to that seen
for field trial 3, but it was not statistically significant
(Figure 1; Table 2).
DISCUSSION
It is clear from our studies that the administration of
probiotics via the drinking water had beneficial effects on
broiler performance. In the field trials, probiotic treatment
significantly improved feed conversion. In each field trial
total final BW was increased by supplemental probiotics,
ranging from 0.74 to 1.64%. Mortality was reduced by
the addition of probiotics to the drinking water. In the
controlled trial there was a marked decrease in mortality
after probiotic administration, which was associated with
a 4.87% increase in total final BW. Unlike in the field trials,
feed conversion was not influenced in the controlled trial,
which may relate to the favorable feed conversion rates
in the latter trial. When the data for total final BW and
mortality were pooled for the 4 experiments, the percent-
age increase in weight and the percentage decrease in
2b 214.3 +67.6 +31.6
2c 214.3 +9.63 +4.49
3a 175.5 +18.9 +10.8
3b 175.5 +27.3 +15.5
4 183.7 +19.7 +10.7
5 168.4 +9.46 +5.62
1
Production number was calculated as follows: kilograms of growth
per day * (100% − mortality) / Feed conversion * 100 (Voeten, 1974).
2
Production numbers could not be calculated for all studies applying
Lactobacillus preparations because mortality was not always mentioned.
Details as to reference, number of animals per treatment, breed, probiotic
description, daily dosage, and administration route are as follows: 1,
Jin et al. (1998a), 6 cages * 10 chicks, Arbor Acres, Lactobacillus acidophilus
(1a) or a mixture of 12 Lactobacillus strains (1b), 1-2 × 106 cfu/g diet,
feed; 2, Jin et al. (1998b), 10 cages * 50 chicks, Arbor Acres, Lactobacillus
casei, 0.5-1.0 * 106 cfu/g diet (2a) or 1-2 * 106 cfu/g diet (2b) or 2-4 * 106
cfu/g diet (2c), feed; 3; Jin et al. (2000), 5 cages * 12 chicks, Arbor Acres,
Lactobacillus acidophilus (3a) or a mixture of 12 Lactobacillus strains (3b),
1-2 × 106 cfu/g diet, feed; 4, Watkins and Kratzer (1984), 2 cages * 50
chicks, not mentioned, undefined Lactobacillus culture, 2 × 109 cfu/chick,
water; 5, Zulkifli et al. (2000), 12 cages * 10 chicks, Shaver × Shaver and
Hubbard × Hubbard, mixture of 12 Lactobacillus strains, 1-2 × 106 cfu/
g of diet, feed.
mortality were found to be at least borderline statistically
significant (P = 0.065 and P = 0.038). An index of produc-
tivity is the so-called production number, which equals
kilograms of growth per day * (100% − mortality) / Feed
conversion ratio * 100 (Voeten, 1974). Table 3 illustrates
that the production numbers for our experiments were
higher than those for the studies reported earlier by oth-
ers. There is a tendency for probiotic treatment to be less
effective when the production number is high. In a study
with veal calves we also noted that growth performance
of the control group was negatively associated with the
magnitude of the effect of probiotics (Timmerman et al.,
2005). Differences in the administration of probiotics
might be another factor affecting efficacy. Administration
of probiotics in the drinking water (Watkins and Kratzer,
1984; present trials) generally resulted in a lower increase
of average daily gain when compared with studies with
probiotic administration via the feed (Yeo and Kim, 1997;
Jin et al., 1998a,b, 2000; Abdulrahim et al., 1999; Zulkifli
et al., 2000; Kalavathy et al., 2003). Another determinant
of probiotic efficacy may be the timing of administration.
During early life, colonization patterns are instable and
chicks are then susceptible to environmental pathogens.
Initial colonization is of great importance to the host be-
cause the bacteria can modulate expression of genes in
epithelial cells (Hooper et al., 2001), thus creating a favor-
able habitat for themselves. These pioneering bacteria will
1388 TIMMERMAN ET AL.
probably reside in the intestine permanently and deter-
mine the colonisation pattern of bacteria introduced later
in life (Ducluzeau, 1993). The primary colonizers are
therefore relevant to the final composition of the perma-
nent flora in full-grown chickens. Prebiotic compounds
supplied early in life may prove beneficial in aiding per-
manent colonization of the probiotic strains administered,
beneficial indigenous microbes, or both.
Overall, probiotic treatment induced a clear reduction
in mortality. Further research is required to study under-
lying mechanisms and to evaluate the economic impact
of the use of probiotics in broilers. Further experiments
with the improved CSPB, as applied in field trial 3, are
required to test whether this less expensive probiotic is
as effective as freeze-dried preparations, which can be
applied via the feed.
ACKNOWLEDGMENTS
The research was supported in part by the Dutch Minis-
try of Economic Affairs (SENTER). The authors wish to
thank R. Blankenstein, L. Mulder, and C. Warmerdam
for technical assistance.
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