Control of Boophilus annulatus (Acari: Ixodidae) on Cattle Using
Injectable Microspheres Containing Ivermectin
J. ALLEN MILLER,1RONALD B. DAVEY,2DELBERT D. OEHLER,1J. MATHEWS POUND,1
JOHN E. GEORGE,1AND ELMER H. AHRENS2
J. Econ. Entomol. 92(5): 1142Ð1146 (1999)
cattle tick, Boophilus annulatus (Say), was tested on 2 groups of 6 Hereford heifers held on separate
7-ha, tick-infested, buffel grass pastures. Cattle in one pasture were injected subcutaneously in the
neck with a controlled-release microsphere formulation of ivermectin at the rate of 2.4 mg AI/kg
body weight; the other group was injected with carrier only. Beginning 4 wk after injection and
on any of the treated cattle, whereas large numbers of engorged ticks were found on the untreated
controls During this period. a few ticks were recovered from untreated sentinel animals placed in
from 11Ð12 wk or 14Ð15 wk. Large numbers of B. annulatus ticks were found on untreated sentinel
habitat were approximately equal, the treated cattle gained an average of 77 kg compared with an
average of 42 kg for the control group. This technology offers a possible alternative to the current
ofÞcial program of dipping and vacating pastures for eradication of Boophilus sp. infestations from
the quarantine zone in southern Texas. Larger scale testing is needed to determine the potential of
the injectable microsphere formulation and to optimize its use in eradication or control strategies.
Boophilus annulatus, ivermectin, injectable microspheres
THE DISCOVERY, DEVELOPMENT, and registration of aver-
mectins, such as ivermectin (Campbell 1989), dor-
amectin (Vercruysse 1993), moxidectin (Webb et al.
1991, Scholl et al. 1992, Miller et al. 1994), and epri-
nomectin (Shoop et al. 1996) as a new class of endec-
tocides (agents for the control of endo-and ectopara-
sites) provide important new tools to control and
manage livestock pests. Not only are the avermectins
efÞcacious against a broad spectrum of livestock pests
(Drummond 1985, Lasota and Dybas 1991), but they
are effective at extremely low dosages when com-
pared with other chemistries. Although numerous
studies have been conducted to characterize the ef-
Þcacy of these compounds against various pests, rel-
atively few investigators have tried to exploit their
unique characteristics by developing novel delivery
systems. Miller et al. (1983) described a sustained
its efÞcacy against larval horn ßies, Hematobia irritans
(L.), and ticks. Boyce et al. (1992) demonstrated that
free-ranging bighorn sheep. The potential for the de-
livery of ivermectin in a sustained release bolus was
explored by Drummond et al. (1981) and Soll et al.
Miller et al. (1998) described a bioabsorbable, in-
jectable microsphere (IMS) formulation containing
ivermectin in poly(lactide-co-glycolide) copolymer
(PLA/PGA) for long-lasting delivery of the drug for
control of livestock pests. Of several possible applica-
tions discussed, they speculated that the injectable
microsphere formulation might be useful and eco-
nomically advantageous for control of the cattle tick,
Boophilus annulatus (Say), and the southern cattle
along the TexasÐMexico border. This barrier is main-
tained by the Tick Eradication Program of Veterinary
Services of USDAÐAPHIS to prevent the reintroduc-
tion of these ticks into the United States from Mexico.
Standard procedures for the eradication of an out-
break of cattle ticks require systematic dipping in
for at least 6 mo, or vacating the premises of all live-
make it difÞcult to gather 100% of the cattle for each
treatment. In addition to being costly, the pasture
This article reports the results of research only. Mention of a
proprietary product does not constitute an endorsement or a recom-
mendation by UDSA for its use.
In conducting the research described in this report, the investiga-
tors adhered to the “Guide for the Care and Use of Laboratory
Animals,” as promulgated by the Institutional Animal Use and Care
Research Laboratory, Kerrville, TX.
1Research, Education, and Economics, USDAÐARS, Southern
Plains Area, Knipling-Bushland U.S. Livestock Insects Research Lab-
oratory, 2700 Fredericksburg Road, Kerrville, TX 78028Ð9184.
2Research, Education, and Economics, USDAÐARS, Southern
Plains Area, Cattle Fever Tick Research Laboratory, P.O. Box 970,
Mission, TX 78573.
vacation approach to eradication may be ineffective
populations of ticks even in the absence of cattle. The
ability to treat cattle with the IMS formulation and,
thus, to maintain the concentration of ivermectin in
their blood above the threshold required to effect
100% control of ticks for 90 d (or 180 d with 2 injec-
tions), would be a major improvement over the cur-
rently available technology.
The objectives of this study were to determine the
concentration of ivermectin in serum resulting from a
selected IMS formulation and to determine if the for-
mulation could control or eliminate a population of B.
Materials and Methods
Pasture Infestations. Two buffel grass (Cenchurus
ciliaius L.) pastures (7 ha, each designated as the east
and west pastures) at the USDAÐARS Cattle Fever
with the Chimenea strain (F7generation) of B. annu-
latus. The Chimenea strain was established from 44
Guy Smith estate, 45 miles northwest of Laredo, TX,
in Webb County. These were reared according to
techniques described by Davey et al. (1984). To ac-
complish approximately equal infestation levels, a
of the tick into each pasture at selected intervals.
intervals for 3 consecutive weeks. At each infestation,
vials containing 5,000 larvae were attached to the
(2 March), 2 of the animals were placed in each
pasture. This procedure was repeated twice for 2 ad-
ditional groups of 4 heifers, with group 2 and group 3
going into the pastures on 16 March and 23 March,
the west pasture died of unknown causes. An animal
was immediately removed from the east pasture to
group of 2 heifers was infested as described and 1
mid-February to early-April, with each pasture con-
taining 6 Hereford heifers weighing ?265Ð270 kg
each. At 19 d (mid-May) before the scheduled treat-
ment, the heifers in each pasture were examined and
estimated to have approximately equal tick infesta-
tions. Unfortunately, only 11 d before the scheduled
uninfested animal was moved into that pasture.
Formulation of Microspheres. Injectable micro-
spheres containing ivermectin were formulated as
previously described by Miller et al. (1998). The par-
ticular polymers selected for this trial were 2 copol-
ymers of PLA/PGA. The 50:50 copolymer (75,000
mw) and the 65:35 copolymer (100,000 mw) were
obtained from Medisorb Technologies International
L.P., Cincinnati, OH. This particular family of poly-
mers and these particular copolymers were selected
because of their safety, bioabsorbancy, and rate of
degradation in vivo. Tice and Gilley (1985), and ref-
cess for the formulation of injectable controlled-re-
lease microspheres of PLA/PGA copolymers and the
formulation. The ivermectin used in the formulation
(Merck, Rahway, NJ) and determined to exceed 95%
purity by HPLC. The resulting formulation contained
as opposed to hollow microcapsules that contain all
the drug in the center. The diameter of the micro-
spheres ranged from ?25Ð250 ?m, with 95% of the
spheres being 70Ð190 ?m in diameter.
from each pasture was switched to the other to pro-
vide balance in cattle weights and tick infestations.
The 6 animals in the east pasture were then randomly
selected as the treatment group and those in the west
pasture served as controls. The 6 heifers in the east
pasture were each treated by subcutaneous injection,
using a 16-gauge needle, into the prescapular area on
of 2.4 mg (AI)/kg body weight with a total volume of
7 ml. The microspheres used in the treatment were a
blend of half 50:50 copolymer and half 65:35 copoly-
mer containing 30% ivermectin. The dosage for each
animal according to its weight was preloaded into a
Johnson and Johnson, Skillman, NJ) was loaded into
another. Just before injection, the outlets of the 2
syringes were connected and the suspending agent
was ßushed back and forth between syringes to sus-
pasture were given a similar subcutaneous injection
containing only the suspending agent.
Data Collection and Analyses. A pretreatment tick
count was taken from each animal in each pasture to
The tick-count procedure used throughout the study
included counting all female ticks that were ?5.5 mm
long on the entire left side of each animal. Because
ivermectin interferes with feeding and not with at-
tachment (Drummond et al. 1981), only ticks that
reached this level of engorgement were counted. In
addition to the actual count of these engorging fe-
males, the overall infestation level was estimated as
low (50Ð100 ticks), moderate (100Ð300 ticks), and
Before treatment (day 0) and at weekly intervals, 2
each animal by using 13-ml SST Vacutainers (Becton
Dickinson, Franklin Lakes, NJ) until no ivermectin
October 1999MILLER ET AL.: INJECTABLE MICROSPHERES OF IVERMECTIN
was detected in any animal for 2 consecutive bleed-
Miller 1989). The technique enables quantiÞcation of
as little as 2 ppb of ivermectin in 5 ml of serum.
Although not speciÞcally designed as a weight gain
study, all animals were weighed before treatment and
after 16 wk. Because cattle were of uniform size and
breeding and were on similar pastures, their growth
under treated and untreated conditions was of inter-
est. General observations also were made on the con-
dition and behavior of the animals within each group.
To determine if the treatment would control the
ticks on the cattle and also eliminate the free-living
tick population in the pasture, 2 untreated, tick-free
Hereford heifers were introduced as sentinels into
Following the 2-wk period of exposure to infestation,
the sentinels were placed in stanchions for an addi-
ticks were collected and counted.
Data on the efÞcacy of the treatment, as measured
by the relative number of engorged ticks in each
pasture, were analyzed using analysis of variance
(ANOVA) of repeated measurements (SAS Institute
1987), and t-tests were used to compare infestation
levels of sentinels and weight gains of cattle in treat-
ment pasture versus control pasture (SAS Institute
Results and Discussion
Three peaks in concentration of ivermectin in the
serum of cattle were evident as a result of the release
ivermectin on the surface of the beads at the time of
and occurred at 3Ð4 d after injection, similar to that
bell 1989). The 2nd peak of ?65 ppb occurred at 4 wk
after injection and is the result of release of the drug
during the hydrolysis of the 50:50 copolymer compo-
degrade and release its drug load, a 3rd peak of ?63
ppb was detected at ?8 wk after injection.
control of ticks feeding on cattle (Drummond et al.
1981). The conventional injection of Ivomec at 200
for only ?3 wk (Campbell 1989). The microsphere
formulation maintains levels of ?8 ppb for ?11 wk.
17.4 engorging females ticks (?5.5 mm diameter),
engorging females on the treated cattle began to de-
cline immediately after treatment. No engorged ticks
after treatment and throughout the remainder of the
of engorged ticks on the untreated controls through-
weeks 13 and 14. Although such natural declines are
common during the warmest period of the summer
(Davey et al. 1994), we have no explanation of this
observed decline. Particularly high numbers of ticks
were found in the escutheon, neck, dewlap, and ßank
areas. In addition, although all animals (treated and
untreated) had high to moderate populations of
nymphs and adults at the beginning of the study, by
were found on the treated heifers. The population of
nymphs and adults on the control animals, although
to moderate levels and, in time, increased to high
A repeated-measures ANOVA indicated a signiÞ-
cattle treated with the IMS formulation and the con-
0.0001). Because most of the observations on the
treated cattle indicated no engorged ticks, all data
were transformed to log10(n ? 1) for the ANOVA.
of 6 heifers as a result of the injectable microsphere formu-
lation at a dosage of 2.4 mg (AI)/kg body weight.
Mean ? SE concentration of ivermectin in serum
2.4 mg (AI)/kg body weight and on control heifers.
1144JOURNAL OF ECONOMIC ENTOMOLOGY
Vol. 92, no. 5
The data resulting from a 2-wk exposure of a pair of
the treated heifers or the control heifers show the
potential of the technology for reducing the free-
living population. During weeks 7Ð8, 178.5 ? 59.5
engorged female ticks were recovered from each sen-
tinel animal placed in the treated pasture. Over 100
times as many (17,966.0 ? 3,095.0) ticks engorged,
detached, and were collected from each sentinel that
of tick-free heifers was put in the control pastures
during weeks 11Ð12, and again a different pair during
weeks 14Ð15, 10,141.0 ? 1,931.0 and 8,653.5 ? 1,389.5
engorged female ticks detached and were collected
Comparisons of the numbers of ticks recovered from
the treated and the controls for each exposure period
ing weeks 7Ð8 (t ? 5.7, df ? 2, P ? 0.05), weeks 11Ð12
df ? 2, P ? 0.05).
The weights of the cattle in the treated and the
untreated groups at the beginning of the study were
270.5 ? 14.9 and 265.5 ? 17.1 kg, respectively. The
cattle, pastures, and tick habitats were approximately
the group treated with the IMS had an ending weight
(week 16) of 347.5 ? 18.6 kg, whereas the ending
weights of the controls was 307.6 ? 18.7 kg. The gain
for treated heifers (76.7 ? 7.2 kg) was signiÞcantly
(t ? 3. 16, df ? 10, P ? 0.01) greater than that for the
control animals (42.0 ? 6.9 kg) over the test period.
This increased gain may be attributed to both the
the treated animals.
From general observations, the treated cattle ap-
peared healthier, developed a better Þnish, and had
a smoother hair coat than the untreated controls.
The untreated controls had a rough hair coat and
acted more irritable when they were gathered for
tick counts and bleeding. We saw no injection site
reaction or other adverse reaction in any of the
In this small-scale study, the use of the IMS tech-
nology not only eliminated B. annulatus from the
treated cattle, but also eliminated the tick population
have enhanced the efÞcacy against a tick with an
?4-wk life cycle.
The IMS technology has a wide range of appli-
cations. It should have applications for eradication
quarantine zone in particular by offering an alter-
native to the labor-intensive, mandatory dipping
program and the expensive pasture vacation pro-
gram currently used. The cost of the current 180-d
program of dipping in coumaphos every 2 wk can
run as high as $60Ð120 per head. This includes the
cost of gathering of cattle and charging the vats, but
does not include the debilitation to cattle in the
process. We estimate that the cost of the IMS treat-
ment could be as little as $25Ð30 per head, and that
the weight gain resulting from the ivermectin treat-
ment could possibly compensate for the cost of the
Although our study was a Þeld or pasture study, it
was on a very small scale. Tests should be conducted
on a larger scale in a naturally infested setting to
determine the full potential of the technology and to
against Boophilus sp., the IMS formulation may have
applications against both endo- and ectoparasites of
zoo animals, bighorn sheep, elk, red deer, and other
exotic game. In the pet market, the formulation may
be useful for control of endo- and ectopararasites
ranging from heart worms to ticks.
The authors thank Dave Krska, Horacio Bazan, Adolfo
Pena, and Homer Vasquez for their invaluable assistance in
handling of animals and assessment of tick populations. In
addition, we thank Alfred Siebenaler for his efforts in data
analysis and preparation of Þgures.
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Received for publication 21 October 1998; accepted 8 June
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