ZINC PHOSPHIDE DEVELOPMENT FOR RODENT CONTROL
Kasem Tongtavee
A Dissertation
Submitted to the Graduate College of Bowling Green State University in partial fulfillment of the requirements for the degree of
DOCTOR OF PHILOSOPHY
December 1978 14
© 1979
KASEM TONGTAVEE
ALL RIGHTS RESERVED ii
ABSTRACT
Adult lab and wild rats (Rattus norvégiens) were maintained under laboratory conditions for the investigation of the bait shyness phenomena, the evaluation of modified forms of zinc phosphide, and the efficacy testing of various bait formulations. One zinc phosphide formulation was selected for a field trial.
Wild rats, but not lab rats, developed bait shyness specific to zinc phosphide following ingestion of sub-lethal doses, but not to the bait material. Some wild, bait-shy rats retained memory for zinc phos phide for at least 60 days.
In food preference studies the rats were given the choice of EPA- standard food and a test food. Only red pepper (at 0.16%) increased palatability of EPA-standard food for both lab and wild rats. Bait combinations A (consisting of ground barley and rolled oats), B (consist ing of ground wheat and rolled oats), and E (consisting of ground barley, ground corn, and rolled oats) were preferred to EPA-standard food by lab but not by wild rats.
Experimental forms of Zn3P2 were mixed with EPA-standard food at 1% and tested on individual lab and wild rats in choice and non-choice regimens. The addition of buffering substances to Zn3P2 did not signifi cantly improve the toxic bait consumption. Most of the special formula tions provided 100% mortality to lab rats in choice and non-choice feeding situations, but only one formulation gave 100% mortality to wild rats.
Seven microencapsulated zinc phosphide formulations provided complete mortality to lab rats in choice tests. Only one of these gave high mortal ity (80%) to wild rats. r in
Four selected zinc phosphide formulations were tested on colonies of wild rats (n=5) in choice tests. Two microencapsulated formulations gave complete mortality to the test animals, and one of these was selected for a field trial. It was mixed with hog food at 1% for baiting Norway rats and house mice outside a hog barn. While this bait was well accepted by these rodents, the population was not reduced significantly because of immigration from surrounding environments.
This study indicated that some wild Norway rats did develop shyness to zinc phosphide as a rodenticide. Special forms of zinc phosphide and microencapsulation improved the efficacy of this rodenticide. Additional development efforts are desirable. • • . ' iv
ACKNOWLEDGEMENTS
I wish to express my thanks to my advisor, Dr. William B. Jackson,
for his helpful suggestions and support throughout this study. I also
wish to express my gratitude to the members of my doctoral committee,
Drs. Robert C. Graves, Francis C. Rabalais, Stephen H. Vessey, John
P. Scott, and Thomas B. Cobb, for their helpful comments and suggestions.
I am very grateful to Drs. Michael W. Fall, Peter J. Savarie, Ray T.
Sterner, Melvyn V. Garrison, of the U.S. Fish and Wildlife Service^ and
John R. Beck, of Biological Environmental Consultant Services, for their helpful comments and suggestions.
I wish to acknowledge the help of A.D. Ashton, who made the facili
ties of the Fernside Laboratory of Bowling Green State University avail
able to me, and the members of the laboratory staff, particularly John
McCumber, Kevin Walsh, Leslie Spooner, Steve Spaulding, Randy Miller,
Kathy Richards, Mike Paessun, Hailu Kassa, and Manfred Temme. I also wish to thank the farm operator, Jim Allion, who allowed me to run a
field trial at his hog farm near Waterville, Ohio. I am very grateful to Ms. Donna L. Dacus, for her job of typing and her care in preventing many errors, and Ms. Rochana Junkasem, for her help in drawing some figures.
My participation in this doctoral program was made possible by the award of a Rockefeller fellowship, and I appreciate this assistance from the Foundation. In addition, Hooker Chemicals and Plastics Corporation provided technical materials and formulations for testing and evaluation.
I am especially grateful for the efforts of A.M. Foster and M.F. Loree of their development staff. I also wish to express my gratitude to the
Thai government for providing my leave of absence so that these studies could be completed.
Lastly, I am deeply grateful to my wife, Vanida Tongtavee, for her understanding and encouragement. vi
TABLE OF CONTENTS
Page
INTRODUCTION 1
Physicochemical Properties and Mode of Action ...... 1
Toxicity and Hazards ...... 4
Stability of Zinc Phosphide...... 7
The Use of Zinc Phosphide...... 8
Food Preferences and Bait Shyness of Rats...... 10
Proposal of Study ...... 17
METHODS AND MATERIALS ......
Experiment 1. Determination of the Phenomenon of Bait Shyness 19
Observation of the behavior of rats in responding to zinc phosphide bait...... 24
Individual lab rats ...... 24
Individual wild Norway rats ...... 25
Determination of types of shyness ...... 25
Determination of retention of memory for bait shyness of wild rats...... 26
Determination of responses of wild rats in social groups to toxic food...... 26
Experiment 2. Investigation of Ways to Increase the Efficacy of Zinc Phosphide ...... 28
Increasing bait palatability ...... 28
Improving zinc phosphide formulations...... 29
Laboratory tests ...... 29
Experiment 3. Field Tests ...... 32
Pre-treatment census...... 33
Treatment ...... 36
Post-treatment census ...... 36 vii
Page
Evaluation of the Efficacy of Zinc Phosphide ...... 36
Additional data for evaluating the efficacy of zinc phosphide...... 37
Use of tracking boards ...... 37
Trapping ...... 37
RESULTS AND DISCUSSION ...... 39
Bait Shyness Phenomena ...... 39
Behaviors of rats toward zinc phosphide baits ...... 39
Nature of bait shyness ...... 57
Retention of memory of bait shyness ...... 60
Social behavior towards bait shyness ...... 62
Improving Toxic Baits to Increase the Efficacy of Zinc Phosphide ...... 69
Bait developments...... 69
Using buffering substances ...... 81
Physical characteristicso f zinc phosphide ...... 85
Improvement of zinc phosphide microencapsulation .... 88
Tests on colonies of wild rats ...... 91
Field Test...... 91
SUMMARY ...... 103
ANNOTATED BIBLIOGRAPHY 105 vili
LIST OF TABLES
Table Title Page
I Oral toxicity of zinc phosphide to various animals. Dose responses are expressed in terms of lethal dose (LD, LD50 LDgg), approximate lethal dose (ALD), or lethal concentra tion (LC50). 5
II The use of zinc phosphide baits for controlling small . rodents in U.S.A. 11
III Zinc phosphide baits used for small rodent control in foreign countries. 12
IV U.S. EPA registered zinc phosphide formulations for commensal rodents. 13
V Experimental zinc phosphide formulations with buffering substances added. ‘31
VI Non-choice tests on rats (R. norvegicus) with Zn3P2 baits. 40
VII Feeding behavior patterns of lab rats in 10-min exposures to ground Purina chow. 42 vin Feeding behavior patterns of lab rats (same rats in Table VII) in 10-min exposures to ZnqPo 0.10% in EPA standard food. 43
IX Feeding behavior patterns of lab rats (same rats in Table VII) in 10-min exposures to Zn3P2atl% in EPA-standard food. 44
X Consumptions by lab rats in three 10-minute experiments in non-choice situations. 48
XI Feeding behavior patterns of wild rats in 30-min expo sures to ground Purina chow. 49
XII Feeding behavior patterns of wild rats (same rats in Table XI) in 30-min exposures to Zn3P2 at 0.10% in EPA standard food. 51
XIII Comparison of feeding behavior and placebo and toxic bait consumption of wild rats (Rattus norvegicus) in three 30-min experiments in non-choice situations. 52
XIV Feeding behavior patterns of wild rats (same rats in Table XI) in 30-min exposures to Zn3P2 at 1% in fiPA standard food. 53 Table Title Page
XV Tests of shyness to wild rats (Rattus norvegicus) In a non-choice situation to different kinds of baits. 59
XVI Retention of memory for bait shyness of wild rats (Rattus norvegicus) to zinc phosphide baits after sur viving a sublethal dose. 61
XVII Pattern of feeding behavior to a colony (n=6) of wild rats in 60-minute exposure to ground chow. 64
XVIII Pattern of feeding behavior (n=3) of a colony of wild rats that survived a sublethal dose of ZrçPj at 0.10% in EPA for six days in exposure to Zng at 1% in EPA in 60-min observations. 67
XIX Consumptions of EPA standard baits modified by addition of enhancers by lab rats in choice tests, five animals per test group. 71
XX Average 4-day food consumption of wild rats (R. norveg icus) in choice tests using experimental baits and EPA standard baits, and EPA + enhancers and EPA standard baits, N=5. 75
XXI Bait combinations tested on lab rats in choice feeding with EPA standard food (N=5). 77
XXII Choice tests on wild rats (Rattus norvegicus) with ZngP2 formulations at 1% in EPA standard food and EPA placebo bait, n=5. 79
XXIII Choice tests on wild rats (Rattus norvegicus) and wild house mice (Mus musculus) with Zn3P2 (with different toxicant formulations) at 1% in EPA standard food and EPA placebo baits, n=5. 80
XXIV Non-choice tests using lab rats (Rattus norvegicus) exposed to EPA standard food containing 1% experimental zinc phosphide combined with buffering substances. 82
XXV Choice tests using groups of lab rats (n=5) exposed to EPA baits containing 1% experimental zinc phosphide formulations and ground Purina chow. 83
XXVI Choice tests using lab rats exposed to EPA baits con taining 1% experimental zinc phosphide formulations and EPA placebo baits. 84 X
Table Title Page
XXVII Non-choice tests using rats (Rattus norvegicus) and mice (Mus musculus) exposed to EPA standard food containing 1% experimental zinc phosphide formulations, n=5. 86
XXVIII Choice tests on lab rats with different Zn3P2 formula tions at 1% in EPA and EPA placebo baits, n=5. 87
XXIX Summary of choice tests using lab rats exposed to EPA standard food containing 1% experimental microencapsu lated zinc phosphide, n=5. 90
XXX Choice tests on colonies of wild rats (n=5) with experi mental zinc phosphide at 1% in EPA and EPA placebo baits. 92
XXXI Summary of bait consumption in field test. 94 LIST OF FIGURES
Figure Title Page
1 (A) Laboratory rats were kept individually in aquaria (75 cm x 30 cm x 27.5 cm) covered with 1-cm wire-mesh in a wooden frame. (B) Interior view of a test room (4.2 m x 3.2 m x 2.8 m) used for observation of rat behavior in responding to zinc phosphide bait. (C) Laboratory set-up with individual cages (25 cm x 20 cm x 17.5 cm) (left), torsion balance (middle), and hy- grothermograph (right). (D) A metal tank (0.8 m x 2.4 m x 0.6 m) used for testing a colony of wild rats with a zinc phosphide bait. 21
2 Wild Norway rats which were individually marked with cloth tape around the tails at different positions. 27
3 Locations of bait stations in pre- and post-treatment censuses, and in treatment period during a field trial for the efficacy of microencapsulated zinc phosphide 30-D at a hog farm near Waterville, Ohio. 35
4 A healthy rat stands on a jar with ease. 47
5 A lab rat that survived a sublethal dose of Zn3P2 at 0.10% in EPA-standard food exhibited muscular spasms before death after consuming Zn3P2 at 1% in EPA-standard food in subsequent exposure. 47
6 Bait consumption by outside populations of rats and mice at a hog farm near Waterville during 16 July - 8 August 1978. 96
7 Rat and mouse tracks on tracking boards (expressed in terms of index numbers) found during the efficacy field trial of Zn3P2 30-D. 98 INTRODUCTION
Zinc phosphide was first used as a rodenticide in 1911-12 in Italy to control field rodents. After that its use gradually increased and by 1930 it was widely used in several Italian cities (Schoof, 1970) .
It occasionally was sold as rat poison in England in the 1930s and until
World War II, when it was broadly used as an effective rodenticide
(Robertson et al., 1945). Zinc phosphide came into use in the United
States shortly after its application in England, and it was used as a substitute for thallium sulfate during World War II (Schoof, 1970).
However, it was not fully developed because of the wartime introduction of compound 1080, which proved to be more effective in controlling rats
(Bentley et al., 1961), and the post-war popularity of anticoagulants.
In recent years, interest in zinc phosphide has increased because it is less hazardous to mem and his domestic animals than memy other acute rodenticides, such as compound 1080, strychnine, and thallium sulfate
(Hood, 1972).
Physicochemical Properties and Mode of Action ■ . 7 , Zinc phosphide (Zn3P2) is a heavy, dark gray, crystalline powder, which is decomposed rapidly by dilute acid or very slowly by water to liberate phosphine gas, which is very toxic and has a garlic-like odor.
It is insoluble in water and alcohol but is slightly soluble in alkalis and oils (Schoof, 1970). This compound decomposes slowly and gives off phosphine gas in contact with air (Elmore and Roth, 1943) and moisture
(Hilton and Robison, 1972).
Commercial zinc phosphide has a range of particle sizes. About 99.5% of this formulation can pass through a sieve of 100 mesh (<150 microns); 3
zinc phosphide particles were found in the livers of rats obtaining lower
doses, they concluded that Zn3P2 particles, not phosphine, directly
caused damage to liver tissues and probably death, if it occurred more
than 24 hours post-ingestion. This is corroborated by unpublished ob
servations (John R. Beck, personal communication). However, I conclude
the major cause of death is phosphine gas absorbed into the blood stream
from the duodenum and its acute effect on the liver, brain, and other organs.
Inhalation by man of phosphine causes restlessness, which soon is
followed by shaking and fatigue, nausea, gastric pain, vomiting and dia rrhea, thirst, headache, dizziness, and oppression in the chest. The patient may have difficulties in breathing and may be coughing with a green-fluorescent sputum. Finally he becomes sleepy and comatose; con vulsions occur shortly before death. Death may be delayed for several days, or it may happen quite suddenly. Continued exposure to subtoxic concentrations of phosphine leads, to chronic poisoning characterized by anemia, bronchitis, gastrointestinal disturbances, dental necrosis, and it may be associated with disturbances of vision, speech, and motor func tions (von Oettingen, 1952).
The inhalation of zinc phosphide dust was followed several hours later by vomiting and diarrhea, rapid and soft pulse, restlessness, and fever, etc., similar to those conditions produced by phosphine (von
Oettingen, 1952). Zinc phosphide also can enter the blood stream through cuts or other breaks in the skin (Anon., 1967b). In fatalities following ingestion of zinc phosphide there was liver, kidney, and heart damage
(Dreisbach, 1963). After repeated doses, kidney damage and hyaline degen 4
eration of the myocardium were observed; livers were damaged, with necro
tic foci usually being in central lobular areas; and lungs had evidence
of serious congestion with blood or exudate in the alveolar spaces (John
son and Voss, 1952).
Toxicity and Hazards
Zinc phosphide is not selective. The LDjq for Norwayr ats is about
40 mg/kg, with a range from 25 to 48 mg/kg (Schoof, 1970). Mammals and
birds accidentally consuming zinc phosphide rodent bait may be killed.
The LD50 for dogs and cats was reported to be about 40 mg/kg; about 50
mg/kg for cows (Hood, 1972). It is highly toxic to birds, the LD50 level
for domestic fowl being between 7 and 17 mg/kg (Robertson et al., 1945).
It is less toxic than chlorinated hydrocarbon pesticides to fish. Cray
fish and shrimp can tolerate concentrations of zinc phosphide in water
from 10 to 50 ppm (Hood, 1972). Toxicity of this compound to animals is
summarized in Table I. Also see Hayes (1975).
While its odor, if not too strong, is attractive to ground squirrels
(Marsh et al., 1970) and other rodents, many believe that its emetic properties and unpleasant odor often make it unattractive to non-target species (Anon., 1976). However, a number of zinc phosphide poisonings were recorded by National Pest Control Association (Anon., 1976) invol ving cattle, pigs, dogs, cats, ducks, and pheasants. Ingram (1945) reported a case in England of a 14 month-old colt dying from eating an unknown amount of zinc phosphide rodent bait.
Induced gastrointestinal irritation following ingestion results in vomiting and diarrhea in most animals (Peoples, 1970). Dogs and cats accidentally consuming zinc phosphide baits often will vomit within five 9
table i
Oral toxicity of zinc phosphide to various animals. Dose res ponses are expressed in terms of lethal dose (LD, LDso, LD90), approximate lethal dose (ALD), or lethal concentration (LC5g).
SPECIES______,______LD LO,g LDSfl ALD LCSO(t) REFERENCES
Rodents:
Norway rat(Rattus nozveglcaa) 40.5 Rudd & Genelly, 1956 Black rat/Rattus zattus) 21.3 Hilton et al., 1972b Black rat(Rattus zattus mindaaensls) 28.0 41.0 West et al., 1973 Grey-bellied rat(R. r. alexandzlnus) 27-47 Helal et al., 1975 White-bellied rat(R. z. fzugivozus) .22-49 Helal et al., 1975 Polynesian rat(Rattus exulans) 23.1 Hilton et al., 1972b Ricefield rat(R. argentiventer) 35.0 Hood, 1972 Grass rat lAzvicanthis niloticus) 2S-40 Helal et al., 1975 Cairo spiny mouse(Acomys cahizlnus) 21-32 Helal et al., 1975 Gerbil(Mezlones huzzianae) 35.0 Fitzwater £ Prakash, Gerbil (Tatera Indies) 35.0 1973 Calif, ground squirrel(Spermphilus beecheyl) 33.1 Hood, 1972 Black-tailed prairie dogfCynomys ludovicianua)10.0 Tletjen, 1976 Northern pocket gopher (Thonootys talpoldea} 6.8 Hood, 1972 Northern pocket gopher(T, t. quadratics) 28.0 Hood, 1972 Kangaroo rat(Cipodoim/s spectabiJis} 8.0 Hood, 1972 Deer mouse(Peromyscvs manlculatus) 42.0 Hood, 1972 Meadow vole(Microtus pennsylvanlcus) 18.0 Hood, 1972 Meadow vole(M. californicusj 15.7 Hood, 1972 Muskrat(Ondatra zibethlca) 29.9 Hood, 1972 Nutria(Myocaster coypus) 5.6 Hood, 1972 Hares:
Jackrabbit fLepua califomicus) 8.25 Evans, 1970 Carnivores:
Dog (Cams famillarls) 40.0 Hood, 1972 Cat(Feiis catua) 40.0 Hood,. 1972 Ungulates:
Cow (pos taurusj 50.0 Hood, 1972 Birds:
Pheasant(Phasianus colchicus) 9.0 Hayne, 1951 Poultry (several species) Blaxland S Gordon, 1945 White-fronted goose (Anser alblfrona) 20-30 7.5 Hood, 1972 , Snow goose(Chen hyperborea) 8.8 Hood, 1972 Mallard duck(Anas platyrhynchos) 35.7 Hood, 1972 Partridge(perdix perdix) 26.7 Hood, 1972 Quail (Lophortyx cali£omica) 13.5 Hood, 1972 Morning dove(Zenaidura macroura) 34.3 Hood, 1972 Sparrow(/»asser dotoesticus) 20-50 Hood, 1972 Red-winged blackbird(Agelaius phoeniceus) 24-178 Hood, 1972 Tricolored blacbird(A. tricolor) 75-316 HOod, 1972 Fishes:
Rainbow trout(Salmo gairdnerii) 0.5 Hood, 1972 Carp(Cyprinus carplo) 0.3 Hood, 1972 Channel catfish(fgtalurus punctatizsl 0.5 Hood, 1972 Black bullhead(f. joelas) 0.4 Hood, 1972 Bluegill (zepomis taacrochirus) 0.8 Hood, 1972 Yellow perch(parea flavescens) 0.6 Hood, 1972 6 minutes and suffer no ill effects. To avoid being hazardous to non-target species, baits may incorporate tartar emetic, which causes vomiting to animals after ingestion (Anon., 1976). However, such an inclusion is not always recommended by veterinarians because of its own side effects.
Rats, however, are unable to vomit (Jackson, 1976).
True secondary hazards may not occur with this rodenticide, since it is not stored in muscle or other tissues of poisoned animals. However, zinc phosphide remains toxic for several days in the gut of dead rats or mice. Therefore, other animals can die after eating recently-killed zinc phosphide-poisoned rodents, if they consume enough of the gut contents
(Anon., 1976). Chitty (1954) reported that observations in the field have shown repeatedly that cats were killed by eating zinc phosphide-poisoned rats. From this viewpoint, Brock (1965) fed gopher snakes (Pituophis catenifer) with mice that had consumed lethal quantities of zinc phosphide and found that the snakes often regurgitated the mice and exhibited no toxic responses.
In field trials for controlling black-tailed prairie dogs with 2% zinc phosphide bait, Tietjen (1976) reported that no non-target species were found sick or dead in the treated areas. Minks, fed daily with
143 grams of ground carcass (except skin and feet) of prairie dogs that had died from zinc phosphide poisoning, received about 13.6 mg/kg of
Zn3P2 from this diet per day and survived the 30-day test period.
Zinc phosphide also is toxic to man. Stephenson (1967) reported that ingestion of 4 to 5 grams of zinc phosphide caused death on two occasions, but a dose of 50 grams has been survived. However, zinc phos phide poisoning should not be confused with yellow phosphorus poisoning 7
(Jackson, 1976), as in a case reporting a 2 year-old boy died five hours later after ingesting a 2.5% phosphorus-containing rat poison (Simon and
Pickering, 1976).
Zinc phosphide has been considered one of the safest acute rodenti- cides; it can be used generally today be pest control operators, government control officials, and farmers for rodent control (Jackson, 1976).It proved safe enough in 1970 to be the first rodenticide granted federal registration for broadcast application over a food crop (Hilton et al.,
1972a,b and Robison and Hilton, 1971).
Stability of Zinc Phosphide
Whether zinc phosphide baits become non-toxic within a few days has been much discussed. This certainly is not true if they are well protected from sunlight and rain or if it has been applied with a non-water-soluble sticker (Anon., 1968). The National Pest Control Association (1967) indicated that zinc phosphide is a stable material and remains poisonous on dry grain baits for months; the deterioration, when being used out doors, is due primarily to physical erosion (the zinc phosphide is washed off the bait materials). Schoof (1970) reported that apples or potatoes, treated with zinc phosphide at 1% by weight and exposed in bait stations or other protected sites, retained 70 to 90% of the toxicant after 90 days; when mixed with wet ground fish toxicity degrades rapidly. Zinc phosphide on oat groats will be broken down if exposed to rain (Hilton et al., 1972), about 60% of the toxicant being degraded in exposure to an inch of rain per day for three consecutive days. Similarly, West et al. (1973) found that polished rice or binlid (broken rice) treated with zinc phosphide at 2% by weight for controlling rats in the Philippines, 8
when exposed to about one inch of rain per day for two to three days, lost
about half of the toxicant. However, even after such losses, the bait
retained enough zinc phosphide to kill rice-field rats. Consequently,
the nature of the bait, the adhesive used, the pH, and the moisture in
the environment all affect the degradability (Guerrant and Niles, 1969).
The Use of Zinc Phosphide
Zinc phosphide has been used effectively as a rodenticide at 0.75
-5% concentrations to control rats and mice in many countries (Anon.,
1975). In India, Kapoor and Khare (1965) used zinc phosphide at 5%
level in grain bait in the rice mill at Saharunpur and exterminated
96% of the rat population. Srivastava (1968) recommended the use of
any type of a whole or crushed grain, such as maize, wheat, or rice,
soaked in water for a few hours and then mixed with zinc phosphide at
2% by weight for controlling rats in India; but in 1972 Deoras recommended
the use of zinc phosphide at 1% level on the same kinds of baits. Zinc
phosphide dusted into rat burrows was effective as a fumigant when mixed
with sulfuric acid-impregnated (20%) clay before application and when
there was enough moisture in the soil (Krishnamurthy and Singh, 1967).
West et al. (1972) used polished rice treated with zinc phosphide
at 1% level for controlling rats in rice fields in Philippines, but they
found that this poison did not appreciably reduce damage done by rats.
They indicated that bait acceptance was a major factor limiting the effec
tiveness of this rodenticide. Twigg (1962) reported that broken rice
treated with zinc phosphide at 2.5% level was effective in controlling marsh rats, Holochilus sciureus berbicensis, which damaged sugarcane in
British Guiana.
/ 9
In the United States zinc phosphide is used at 1% weight in suitable
bait materials, such as cat and dog food, poultry mash, whole grains,
rolled oats, corn meal, or bread crumbs, for house mouse and rat control.
Fresh fruits or vegetables, such as cantaloupe, apples, sweet potatoes,
cut into pieces (about 1/2 inch cubes), can be coated with zinc phosphide
at about 1% by weight for controlling rats and mice (Anon., 1976). Marsh
(1966) suggested using zinc phosphide grain baits at 1 or 2% for control
ling rats in rice fields. Clark (1972) recommended that oat groats treated with zinc phosphide at 1% by weight or 1/2 inch cubes of sweet potatoes dusted with zinc phosphide at about 1.5% by weight be used for cotton rat control in California. Other small rodents, such as meadow mouse (Micro-
tus spp.), kangaroo rat (Dipodomys spp.), and prairie dog (Cynomys spp.), can be controlled with zinc phosphide apple or grain baits at a 1% level
(Ludeman, 1962). It is commonly used against sewer rats (Rattus norvegi cus) at 1 to 2.5% in cereal grains; it can be mixed with ground horse meat or ground fish (Brooks, 1962). While 2% baits are more effective in the proper bait, they usually are less acceptable than the lower concentration forms.
Since the efficiency of zinc phosphide bait will be decreased when the bait is exposed to rain for several days, it has been suggested that the bait should be prepared in paper-wrapped "torpedoes" so that its qualities will be extended (Gratz, 1973). Deoras (1972) reported that
Zinc phosphide bait coated with paraffin as moisture proofing killed many rats in rat campaigns in India. Kusano (1977) developed zinc phosphide bait in a form of a small, irregular cube or pyramid "pellet" (consisting of a mixture of various grain powders and zinc phosphide at 3%) and found 10
that it had high efficacy for controlling the Japanese field voles
(Microtus nontebelli) in agricultural areas. Other reports of zinc
phosphide controlling rodents are presented in Tables II, III, and IV.
Since it had been reported that the rats did not get a lethal dose
from eating the poisoned grain baits distributed in Hawaiian sugarcane
fields by aerial baiting, a tablet bait was developed. A single zinc
phosphide tablet(consisting of corn and oat groats as a bait carrier
and weighing approximately 300 mg each and containing 4% Zn3P2) provided
100% mortality for Polynesian and wild Norway rats under laboratory con
ditions (M.V. Garrison, personal communication).
Another approach of using zinc phosphide as a rodenticide is the
application of this compound as a tracking powder (e.g., place in runways
and burrows of rodents). The advantage of using tracking powder is that
in grooming the rats will ingest the poison adhering to their fur and feet.
Furthermore, its use is possible against the population which develops bait shyness after surviving a sublethal dose of zinc phosphide bait.
Marsh (1973) tested zinc phosphide tracking powder at a 5 or 10% level on house mice (Mus musculus) under laboratory conditions and found that it caused 100% mortality to the mice. It is now EPA-registered for use.
Food Preferences and Bait Shyness of Rats
While rats generally feed on a wide variety of foods, rats in dif ferent habitats may show differences in food preferences. For instance, rats in cities may prefer garbage to grains; whereas, rats in agricultural fields may prefer food grown in those particular areas. Brooks and
Bowerman (1973) reported that wild Norway rats trapped from a disposal TABLE II
The use of zinc phosphide baits for controlling small rodents in U.S.A.
Controlled Species Bait Component Adhesive % Zn3P2 Amount of poisoned References in bait bait used in field
1) Ground squirrel a) Oat groats Petrolatum 0.6 1 pound/acre Storer, 1958; (Spermophilus beecheyi) b) Whole barley or Storer and Jameson, 1965 whole oats Petrolatum 1.0 1 pound/acre 2) Ground squirrel Whole barley (re- Lecithin-mineral 1.0 1 pound/acre Dana, 1962 (S. beecheyi) cleaned) oil 3) Ground squirrel Oat groats 1.0 1 pound/acre for Clark, 1975 (S. beecheyi) spot baiting 2.0 6 pounds/acre for broadcast baiting 4) Rats and house mice a) all fish (cat and 1.0 HPCA, 1967 (Rattus norvegicus dog food) 89 parts #17-67 (Anon., 1967b) and Hus musculus) by weight and corn meal 10 parts by wt. b) Poultry mash 84 Com oil or bacon 1.0 parts by weight grease IS parts by weight 5) Meadow mouse Cracked corn, oat Colored with lamp 0.5-1 .0 Rudd & Genelly, 1956 (Microtus spp.) groats« or wheat black or with methylgreen 6) Cotton rats a) oat groats lecithin-mineral 1.0 for hand baiting Clark, 1972 (Sigmodon hispidus) Oil 2.0 for broadcast bait. b) cubed sweet Vegetable oil 1.5 potatoes 7) Black-tailed prairie Oats Corn oil 2.0 Tietjen, 1976 dogs (Cynomys ludovicianus) TABLE III
Zinc phosphide baits used for small rodent control in foreign countries.
Species Bait Components Adhesive Additive % Zn3P2 Countries References in bait
1) Norway rats Maize meal 65% Corn oil 5% Sugar 5% 2.5, 5.0 England Rennison et al., 1968 (Rattus norvegicus) Rolled oats 25% 2) Norway rats Medium oatmeal Com oil 5% Sugar 5% 2.5, 5.0 England Rennison et al., 1968 (R. norvegicus) 3) Norway rats Damp coarse oatmeal (R. norvegicus) (Pinhead oatmeal: Com oil 5% Sugar 5% 2.5, 5.0 England Rennison et al.. 1968 water 2:1) 1.0, 2.5 Rennison, 1976 4) Norway rats Sausage rusk Cora oil 5% Sugar 5% 2.5, 5.0 England Rennison et al., 1968 (R. norvegicus) 5) Wild rats Medium oatmeal Com oil 5% Sugar 5% 0.1, 0.2 England Greaves, 1966 (R. norvegicus) 0.4, 0.5 0.6, 1.0 2.5, 5.0 6) Mice Medium oatmeal Sugar 10% 1.0, 2.0 England Cornwell, 1971 (Mus musculus) 4.0 7) Marsh rat Broken rice plus 2.5 British Twigg, 1962 (Holochilus sciu maize Guiana reus berbicensis) 8). RicefieId rats Polished rice Coconut oil 1.0 Philippines West et al., 1972 (R. rattus Coconut chunks l-cm^ 0.5% 1.0 West et al., 1972 mindanensis) 9) Bandicota bengalen- Cereal; (i.e, wheat, Vegetable oil 2.0 India Srivastava, 1968 sis bengalensis rice, maize, etc.) 10) Bandicota Bice flour 80% plus Groundnut oil 2.0 India Durairaj and Guruprasad bengalensis roasted groundnut 10% 10% 11) Field rodents (i-e., Broken rice 95.5% Refined cottoi 2.5 Pakistan Greaves et al., 1977 B. bengalensis, seed oil 2% Mllardia meltada, Mus spp., Nesokia indica, Rattus rat tus, Tatera indica)
12) Mus musculus spici- Com and oat Com oil 3.0 Switzerland Hamar, 1976 legus, Apodemus kernels sglvaticus, A. agrarius, Rattus norvegicus
K> 13 TADLE IV
U»S. EPA-reglster«d zinc phosphide formulations for conmonaal rodents.* (*Data obtained from documents supplied by U.S. EPA to Or. William B. Jackson, Director, Environmental studios. Dowling Green State Univ.)
EPA Regis- Manufacturing Company Trade Name Activa For Control of Bait Base Additive Substances Percent tratlon Ingré Recommended Recomnendod ZnjJfc in Number dient » the bMt
355-1 American Fluoride RUMETAN 94 House mice(Mus 1) Rolled oats Com oil. cotton seed oil, light 1 Corp.■ New York. NY musculus) and 2) Hamburger. mineral oil or bacon fat Rats (genus canned sal Rattus) mon, etc. 3) Sliced banana, apple*, etc.
255-58 As above RUMETAN SO As above As above As above 1 255-86 As above Sine phosphide 74.7 As above Soiled oats, Bacon grease or icom oil 1 bread crumbs, diced fresh coconut, fish, ground neat, etc.
1691-82 Chemical Compounding Zinc phosphide 74.7 As above As above As above 1 Corp. Jersey City, NJ
4887-68 American Fluoride Co. RUMETAN 94 Norway rats 1) Grains Corn oil, cotton seed oil, light 1(house) New York. NY (R. norvegicus/ ciner&l oil, or bacon fat Roof Rats. .. (R. rattus/. and House mice. 2) Meat Glycerine oil 2(dunp) (Jfus musculus/ 6704-36 Pocatello Supply Depot Zinc phosphide 63 As above Ground meat. —— 1 Bureau of Sport Fish Rat Poison fish, canned eries 6 Wildlife dog or cat food, Pocatello. Idaho poultry mash, et&. 6720-115 Southern Mill Creek Zinc phosphide 80 As above Canned dog or 1 Products Co.. Inc. cat food; Com Com oil or melted kitchen 1 Tampa. Florida meal, chicken grease ■ash, bread cr. 7122-30 ArChem Corporation GUASDIAN Zinc 94 As above A ready to use 1 Portsmouth. Ohio phosphide (POO use bait only) 6913-5 Abalene Pest Control ZINCO Rodent 1 As above Cracked corn Corn oil 1 Service. Inc. • Bait (Seedy to use Poughkeepsie. NY bait)
“Ï — 485-31 WINRU Chemical & Sales WINRU 2 White-footed Ready to use Ooapany mice (genus Kansas Ciy. Missouri Perorayscus) Meadow mice. (genus Microtus) Pine mice. (genus Pitymys)
728-51 Pearson fi Company PEARSON’S 2 Cotton rat (ge- Ready to use 2 Mobile. Alabama RAT POISON nus Sigmodor) and field mice (ge nus Peromyscus, Microtus fi Pitytaysi 7122-31 ArChem Corporation GUARDIAN 1 Meadow mice A ready to use 1 Portsmouth, Ohio PHOSPHOR-GRITS {Microtus bait Deer mice(Pero- ayscus spp.) Pine mice (Pitymys spp) House Mice(tfus mus culus/ fi Rata Rattus spp.) 7122-82 ArChem Corporation PARA ZINC 1 Rats Rattus Paraffinized- 1 Portsmouth. Ohio spp.) pelletized bait 10646-1 Hawaii Dept. of Agri. 1.88 Polynesian rats Oat groats 1.88 Honolulu. Hawaii (R. exulsns), A ready to use Norway rats (R bait norvegicus/Black rats(R. rattus/ 10741-1 KILLRAT Corporation KILLRAT 1 Rats fi Mice (ge A ready to use 1 Phoenix. Arizona nus Rattus fi JftisJ* bait 11555-1 Int’l 2000. Oklahoma MR. RAT GUARD 1 As above A ready to use bait 1 18710-1 Sanford Chen. Co. NU-KIL 1 As abova A ready to use bait 1 Oklahoma City, OK 12455-16 Bell Laboratories ZP Tracking 10 House mice Ready to use; — Madison. Wisconsin Powder fffus musculus/ sprinkle in patches 14
site in New York preferred sweet rice, millet, peanuts, milo, barley,
sunflower seeds, and oats over corn meal; whereas, those that were trap
ped from a chicken farm preferred millet, sweet rice, and peanuts over
corn meal. Khan (1974) tested the black rat (Rattus rattus) for food
preferences and found that 1) of whole cereals, millet was preferred to
wheat, and both of them to maize, 2) semolina (extracted flour ground
into granules) was preferred to millet flour, maize flour, and wholemeal,
3) the rats showed preference for moist food and for food with groundnut
oil or sugar-cane, 4) groundnut oil was preferred to sugar.
Rats, initially, may select familiar food (Jackson, 1972); however,
they may change from a familiar food to a novel food if the latter is
more palatable (Barnett, 1956). Krishnakumari (1973) conducted experi
ments that indicated early experience had little influence on food prefer
ences of adult Norway rats. -
Rats and mice use cues, such as taste or odor, in learning to avoid
poisoned food by associating their illness from a sublethal dose with
that particular food or bait (Jackson, 1974; Wallace, 1976). While Barnett
et al. (1975) reported that the black rats (Rattus rattus) surviving a
sublethal dose of Zn3P2 bait avoided feeding on millet coated with this
compound on the following days, Heinz (referred to by Gaffrey, 1953)
found the contrary. Rozin (1968) stated that rats tended to avoid the
food eaten earlier if the food had a novel, distinctive taste, was a
poison, and/or was vitamin deficient. Hargrave and Bolles (1971) empha
sized the importance of taste rather than visual and olfactory cues in
establishing bait shyness; Hankins et al. (1973) stated that rats did use olfactory stimuli in the rejection of a toxic food even after a taste 15
aversion had been established.
Barnett et al. (1975) observed that anosmic rats [Rattus rattus,
wftxcn. had theihr nadil sulfate) that had received a sublethal dose of Zn3P2 obviously avoided contact with poisoned food again. They concluded that bait shyness does not depend on the olfactory sense. While a rat has a long-term memory for taste aversion, the strength of aversion was inversely related to the time interval between consumption and illness (Nachman and Jones, 1974). Also indicated was that the intensity of aversion acquired by rats decreased with increasing time in terval between CS (conditioned stimulus, e.g., the taste of an ingested solution) and US (unconditioned stimulus, e.g., an induced illness). Experiments using drugs and X-irradiation have been conducted to study learned taste aversions of lab rats. Garcia and Roelling (1966) reported that lab rats avoided the food they had eaten if they were sub sequently made sick from lithium chloride injection or from X-irradiation. Nachman and Ashe (1973) found that lab rats were highly sensitive to learning a taste aversion with Lid whether administered intraperitoneally, subcutaneously, or by stomach tube. Taste aversion also occurred in lab rats after the injection of apomorphine, an emetic drug (Garcia et al., 1966), and cyclophosphamide, a drug that produces diarrhea (Garcia et al., 1967; Wilcoxon et al., 1971). Also see Dorrance and Gilbert (1977). Furthermore, rats tended to associate a given taste with poison not on the strength of the taste but on the magnitude of the novelty of this taste (Kalat and Rozin, 1970; Kalat, 1974). Revasky and Bedarf (1967) treated lab rats with X-irradiation after permitting them to ob tain both a novel and familiar food and later found that rats had aversion 16 only to the novel food. Nachman and Hartley (1975) treated lab rats with various rodenticides (i.e., copper sulfate, sodium fluoroacetate, red squill, warfarin, thallium sulfate, sodium cyanide, and strychnine sulfate) injected intraperitoneally after feeding and found that cyanide, strychnine, and warfarin were ineffective in producing a learned taste aversion to a novel food, whereas copper sulfate, red squill, and sodium fluoroacetate were relatively effective; thallium sulfate produced only a slight aversion. Prakash and Jain (1971) reported that two gerbils (Tatera indica. and Meriones hurrianae) developed aversion to zinc phosphide after one day’s exposure to a sublethal dose. Hana et al. (1975) also found that the Hairy-footed Gerbil (Gerbillus gleadowi) associated the sickness developed from feeding upon a sublethal dose of zinc phosphide to the food mixed with this compound. Other acute rodenticides also have similar disadvantages. Crabtree (1962) reported that rats learned to associate the taste of strychnine with its effects and rejected such poisoned baits on repeated exposure. Those that survived a sublethal dose of red squill developed bait shyness that persisted for four to six weeks. Compound 1080 induced the development of bait shyness of white-footed mice (Tevis, 1956). Howard (1959) also reported that laboratory rats first fed a sublethal dose of compound 1080 learned to reject any bait offered that contained a lethal dose of this compound. Gaines and Haynes (1952) reported that wild Norway rats retained bait shyness to ANTU at 2% level for three months after having survived a sub lethal dose. Rzoska (1953) found that all lab rats that survived a near fatal dose of arsenic in bread paste refused an identical poisoned bait 17 after 46 days; two weeks later 11 out of 12 animals still rejected it. In another experiment, he reported that lab rats having experience with red squill powder in bread paste rejected barium carbonate in the same base a month later, thus indicating that the rats did not develop shyness to only the toxicants but also to the bait materials. Robbins (1977) repor ted that taste aversion to a flavor associated with illness induced by lithium chloride also occurs in deer mice (Peromyscus maniculatus) and persists indefinitely if non-toxic fluid is simultaneously available. The development of bait shyness by rats seems to be complicated; there are variations depending upon the nature of the bait materials, the poison, and individual behavior (Rzoska^ 1953). Zinc phosphide, while an effective rodenticide for rats and mice, has the disadvantage that this phenomenon may be quickly developed (Marsh et al., 1970), although consi derable variability may exist. Proposal of Study Zinc phosphide has been used for rodent control in many countries for many years; however, the efficacy of this compound in the field was not always satisfactory. Many factors (e.g., bait acceptance, bait shyness) may have been involved in reducing the effectiveness of this rodenticide. Though often mentioned, little attempt has been made to analyze these phenomena more closely. The purpose of this study was to 1) investigate the behaviors of rats responding to a Zn3P2 bait before and after exposure to a sublethal dose, 2) determine the retention of bait shyness in Norway rats (Rattus norvegicus), 3) consider the types of shyness (whether it is shyness of a rodenticide or the avoidance of a bait material), and 4) evaluate the 18 alternatives. . The improvement of toxic baits to increase the efficacy of zinc phos phide was investigated in the laboratory by 1) the addition of buffering substances to the baits to slow down the hydrolysis of zinc phosphide in the animal's stomach, 2) the use of encapsulation for masking the taste and odor of this chemical and delaying the digestive action so that the animals are unable to associate their illness with the toxic food, 3) the manipulation of physical characteristics of the zinc phosphide to reduce the gustatory response so that the rats would eat more toxic food, 4) the addition of bait enhancers (e.g., salt, red pepper, brown sugar, etc.) to increase the palatability of the food. Different combinations of grains (i.e., com, barley, soybean) were tested to improve the palatability of the baits. Zinc phosphide bait combinations that proved most effective under laboratory conditions were selected for evaluation in free-living colonies and under field conditions. 19 METHODS AND MATERIALS Most of the experiments were conducted at Fernside Laboratory and the Department of Biological Sciences, Bowling Green State University. Adult laboratory rats (Spartan strain of Sprague-Dawley) were obtained from Spartan Research Animals, Inc., Haslett, MI for use in the tests of bait shyness, food preference, and efficacy of different zinc phosphide baits. Adult wild Norway rats of both sexes also were used in some experiments. They were trapped in the Bowling Green area and kept in separate cages for at least one week before the beginning of each experi ment. Most of the test rats weighed at least 150 grams. Adult wild house mice (Mus musculus) also were obtained locally and used in some experiments. Their body weights were between 12 and 28 grams. Any ani mals that showed symptoms of sickness or had wounds and pregnant females were discarded. In order to observe the responses of lab and wild rats to zinc phosphide baits, the test animals were kept individually in covered aquaria (Fig. 1A). A wide-mouth one-gallon jar provided a nesting site. For observations of responses of wild rats in social groups to zinc phosphide baits, the test animals were allowed to freely range in a test room and provided with two wooden nest boxes (Fig.IB). Red fluorescent light was used as the night light to make possible observations of wild rat behavior, since it has been reported that under such conditions ani mals could be trapped, observed, and recovered with little apparent disturbance, reaction, or fear of handling (Fall, 1978). However, it appeared that wild rats kept in aquaria were frightened if the observer were present in the test room; therefore, a curtain with a small observa- 20 Figure 1A. Laboratory rats were kept individually in aquaria (75 cm x 30 cm x 27.5- cm) covered with 1-cm wire-mesh in a wooden frame. Figure IB. Interior view of a test room (4.2 m x 3.2 m x 2.8 m) used for observation of rat behavior in responding to zinc phosphide bait. Figure 1C. Laboratory set-up with individual cages (25 cm x 20 cm x 17.5 cm) (left), Torsion balance (middle), and hygro- thermograph (right). Figure ID. A metal tank (0.8 m x 2.4 m x 0.6 m) used for testing a colony of wild rats with a zinc phosphide bait. Figure 1A Figure 1C Figure 1D 22 vation hole was hung between the rats and the observer to reduce such effects. , For the purpose of testing food preferences and efficacy of different experimental zinc phosphide baits, the test animals were individually caged (Fig. 1C) . A metal cattle-watering tank covered with a wire-mesh and provided with two wooden nest boxes also was used for testing reac- r- tions of colonies of wild rats (n=5) to zinc phosphide baits (Fig. ID). A torsion balance (accurate at 0.1 g) was used for weighing the animals, food, and treated baits. The temperature and relative humidity of the test room at Fernside Laboratory were recorded using a hygrothermo- graph (Fig. 1C). The temperature (of representative 21-day record) was between 17 °C and 28 °C; the relative humidity, between 32 and 58 per cent. The photoperiod was automatically controlled for 12 hours of light and 12 hours of darkness (red-light condition). EPA-standard food was used as the challenge diet in the tests of food preference and in most cases as a bait carrier for experimental zinc phosphide formulations. It was made of 65% ground com, 25% rolled oats, 5% powdered sugar, and 5% corn oil and mixed thoroughly, using a twin- shell bait mixer for about two hours. The additive substances used for the tests of food preferences were salt, red pepper, granulated sugar, brown sugar, monosodium glutamate, garlic powder, and coconut flakes pur chased at local grocery stores. Each of them was mixed thoroughly into EPA standard food in stated concentrations (by weight). Other grain baits similarly were made from different mixtures of ground corn, barley, wheat, and soybean combined with powdered sugar and com oil. Bait X (consisting of 94% barley and 5% vegetable oil) also was made as a bait carrier for 23 zinc phosphide bait shyness tests. All zinc phosphide formulations used in this study were supplied by Hooker Chemical Company. Each was mixed thoroughly with EPA-standard food to make a toxic bait at 1% by weight (or 0.5% in some ejq?eriments). In all cases, test animals were fed Purina lab chow before and after experiments. Water (in bottles or chick fountains) was available at all times. The spilled food was collected from the dropping tray or open floor and added to the respective food dish to obtain an accurate measure of daily consumption. For testing efficacy of zinc phosphide baits under laboratory condi tions, a choice test was performed in which each rat was offered two kinds of food: one a toxic food and the other EPA-standard food (with some exceptions, which are discussed individually) in separate containers for three days. The positions of food bowls were alternated daily to avoid position preferences of the rats affecting the consumption patterns. A non-choice test was carried out in some experiments with each rat offered only the toxic food for three days. Experiment 1. Determination of the Phenomenon of Bait Shyness A sublethal bait was developed by testing three groups (n=5 or 6) of individually caged laboratory rats with zinc phosphide baits at dif ferent concentrations in EPA-standard* food. After two days of pre-test with ground Purina chow and 24-hour food deprivation period, the animals in group I (n=5) were offered zinc phosphide bait at 0.25% for one night; group II (n=5), 0.10% bait for one night; and group III (n=6), 0.10% bait for 20 minutes. Since more than 80% of the rats died in the above experi ment, in the following experiments the rats were presented with, zinc 24 phosphide bait for only 10 minutes after being deprived of food for 24 hours. Those rats that did not ingest 15-30 mg/kg of toxicant were presented with the toxic bait for longer periods until the required dose was obtained. A. Observation of the Behavior of Rats in Responding to Zinc Phosphide Bait. The objective of this study was to determine the occurrence of vari ous behaviors of both lab and wild rats, especially feeding behavior (the rats consuming food) and investigative behavior (the rats sniffing in or around the food bowl), during each minute of the observation period. The occurrences of other behaviors, such as exploring (the rats walking and investigating surrounding environment), grooming, and drinking, also were observed and recorded. A.l. Individual lab rats Five laboratory rats were kept individually in aquaria for two days to habituate to a new environment before the beginning of L . . r' the test. Their responses to the baits were observed for 10 minutes of each test period, and the amounts of bait consumed by each rat also were determined. After being deprived of food for 24 hours, the first bait offered was ground Purina lab chow. This was left until the following day; then the rats were deprived of food for another 24-hour period before being offered zinc phosphide at 0.10% in EPA-standard food in a non-choice situation. The amount of poisoned bait eaten by rats as a sublethal dose was expected to be in a range of 15 to 30 mg/kg. Those rats that did not 25 receive this amount were offered the toxic food for longer periods until the required dose was taken. Symptoms of sickness were twice observed at subsequent two-hour intervals. The rats did not receive any other food until the following day, so they could relate their sickness only to the last food eaten. The rats that survived a sublethal dose of zinc phosphide at 0.10% in EPA were fed ground Purina chow for five days; then they were retested with zinc phosphide at 1% in EPA in a non-choice test situation for one day after being deprived of food for 24 hours. A.2. Individual wild Norway rats Five wild rats were tested with the same procedure (A.l), except that each observation period was extended to 30 minutes. The red light and a curtain also were set up for facilitation of the observations. B. Determination of Types of Shyness Three groups (n=6) of wild Norway rats were used for this study. They were kept in separate cages and fed with ground Purina chow for two days, so that the normal daily consumption of each rat could be determined. Then the rats were deprived of food for 24 hours before being presented with zinc phosphide at 0.1% in EPA for 30 minutes. The amount of poison consumed by each rat was expected to be in a range of 15 to 30 mg/kg. Those rats that did not get enough poison were offered the same poison bait for another two-hour period or even over night. However, the rats did not receive any other food until the following day. The rats that survived a sublethal dose were fed with ground Purina chow for five days. Then the survivors from group I were offered EPA standard food; group II, with zinc phosphide at 1% in EPA standard food; and group 26 III, zinc phosphide at 1% in bait X (consisting of 94% barley and 5% vegetable oil) in a non-choice test for one day. In each case food consumption was measured. C. Determination of the Retention of Memory for Bait Shyness of Wild Rats. Three groups (n=6) of wild Norway rats were used for this study. To obtain the survivors of a sublethal dose of zinc phosphide, the rats were tested with the same procedure explained in l.B. The animals that recovered and ate normally were retested with zinc phosphide at 1% in the same bait material. The survivors from group I were retested in two weeks group II was retested after surviving for one month, and group III was retested after surviving/ for two months. Ground Purina chow was given to the test rats between the presentations of the toxic food. D. Determination of the Responses of Wild Rats in Social Groups to Toxic Food. The purpose of this study was to determine social behaviors (e.g., hierarchy in feeding) of wild Norway rats in responding to zinc phos phide baits. The occurrences of fighting among individuals at times of feeding also were observed and recorded. A group of seven wild rats (one female and six males) were kept in the free-ranging test room (Fig. IB). Each rat was marked individually with a cloth tape around its tail (Fig. 2). Ground Purina chow was pro vided for the rats only at night for about 14 hours in 24-hour period. Dead animals were removed from the test room whenever they were discovered The social behaviors of rats responding to ground Purina chow were 27 Figure 2. Wild Norway rats which were individually marked with cloth tape around the tails at different positions. 28 observed under red light for one hour on the seventh day at time of feeding. After that the rats were transferred to individual cages and fed with ground Purina chow for two days in order to obtain the daily consumption of each rat. The rats were deprived of food for 24 hours before being offered zinc phosphide at 0.10% in EPA-standard food for 30 minutes. Each rat was expected to receive about 15 to 30 mg/kg of poison as a sublethal dose. Those whose consumption was not within this range were offered toxic food until the required dose was taken. After that the rats were fed with ground Purina chow for five more days. The animals that recovered from toxic symptoms were returned to the test room. Two days later they were offered zinc phosphide at 1% in EPA- standard food in a non-choice test for 24 hours. The occurrences of so cial behaviors of rats responding to toxic food were observed and recorded again for one hour at the time of feeding. The amounts of toxic food consumed by rats were recorded after the end of observation and after 24 hours of consumption. Experiment 2. Investigation of Ways to Increase the Efficacy of Zinc Phosphide A. Increasing Bait Palatability Two bait formulations (one EPA-standard food with additives, the other a combination of different grain baits) were tested against the EPA challenge food on lab rats. The additive substances used for this study were salt, red pepper, granulated sugar, brown sugar, monosodium glutamate, garlic powder, and coconut flakes. Different combinations of grain bait included ground corn, ground barley, ground wheat, and ground 29 soybean combined with powdered sugar and corn oil in various concentra tions. The lab rats (n=5) in each group were caged individually and tested with one of the bait formulations. They were offered two kinds of food (one a bait formulation and the other EPA challenge food) in separate food bowls for four days. The positions of the food bowls were alter nated daily to avoid position preference of the rats. Some groups of the rats participated in more than one feeding test, but no repetition within any sequence occurred. Statistical analyses were performed, using t-tests, for comparisons of food preferences. All of the bait formulations preferred to EPA stan dard food by lab rats were selected for similar tests on wild rats. Any formulations that were preferred to EPA food by wild rats were selected again for use as bait carriers of the experimental zinc phosphide for the tests on wild rats. B. Improving Zinc Phosphide Formulations Laboratory tests 1.1. Tests on individual rats Most formulations of experimental zinc phosphide were tested on two groups (n=5) of lab rats, one group for non-choice test and the other group for choice test. In all experiments, the rats were tested for 10-day peri ods, two days in pre-test, three days in test, and five days in a post-test. They were fed with ground Purina chow in pre- and post-test. Any formula tion that was effective against lab rats (providing 100% mortality in a choice test) was selected for the tests on wild rats using the same procedure. 30 The different zinc phosphide formulations used are characterized to the extent possible. In some situations only code designations can be used because of the development of patent information. a. Zinc phosphide combined with buffering substances. Inorganic salts (at 40%) were mixed with Zn3P£ on a ball-mill for 20 minutes. Part of the samples were water washed at 80-90 °F (27-32 °C) for six hours. Eight samples of zinc phosphide were prepared for this study; their characteris tics are indicated in Table V. Each of the samples from number one to six was mixed thoroughly in EPA-standard food at 1% level, while samples number seven and eight were mixed at 1.6% level. In some cases ground Purina chow was used instead of EPA-standard food. b. Physical modifications of technical zinc phosphide. Ten samples of different forms of zinc phosphide were used for this study. Each. was mixed thoroughly with EPA-standard food to make a toxic bait at the 1% level and was tested on two groups (n=5) of lab rats with the same pro cedures as explained in 1.1. Any formulation that was effective against lab rats was selected again for the tests on wild rats and wild mice. . c. Microencapsulations of zinc phosphide. Eighteen samples of microencapsulated zinc phosphide were used for this study. Each sample was thoroughly mixed with EPA-standard food to make a toxic bait contain ing 1% zinc phosphide and tested on a group (n=5) of lab rats in a choice test procedure. Any formulation that was effective against lab rats was selected for the tests on wild rats. 1.2. Tests on colonies of rats . Any formulations of zinc phosphide that were effective against wild rats in the choice tests were selected for the test on a colony (n=5) 31 TABLE V Experimental zinc phosphide formulations with buffering substances added.* Formulation Buffering Substances Percentage Water Wash** 4369-2-1 MgCO3 40 + 4369-21-2 NaHCO3 40 + 4369-21-3 CaHPO3 40 + 4369-21-4 MgCO3 40 - 4369-21-5 NaHCO3 40 - 4369-21-6 i CaHPO3 40 - 4369-21-7 None + 4369-21-8 None ■ - *33.3 gm of the inorganic salts were combined with 5Ogm technical (94%) Zn3P2 **washed at 80-90 °F for 6 hours 32 of wild rats. The rats were fed ground Purina chow for three days in pre-test. Then they were offered a selected zinc phosphide mixed with EPA-standard food at 1% level in two experimental food bowls and EPA- standard food in the other two food bowls for three days. Two of the food bowls (one contained EPA-standard food and the other contained a toxic bait) were placed side by side at the left side of the metal tank, and the other two food bowls (one contained EPA-standard food and the other contained a toxic bait) were placed side by side at the right side of the metal tank. The positions of the two food bowls in each station were alternated daily to avoid position preference by the rats. The survivor rats were fed ground Purina chow five more days for observation. Daily bait consumption and mortality of the rats were recorded. Experiment 3. Field Tests A zinc phosphide formulation that produces 100% mortality to wild rats under laboratory conditions may not achieve control of rats under field conditions, since wild rats tend to avoid new objects and feed on foods familiar to them (Chitty and Shorten, 1946). Thus they may not accept the toxic bait readily. For this reason field trials are necessary. One zinc phosphide formulation that had proved efficacious against wild Norway rats under laboratory conditions was selected for testing at a hog farm located near Waterville, Ohio. This site was infested by Norway rats and house mice, as evidenced by the presence of fresh droppings, burrows, and runways outside the buildings. Holes gnawed through the walls and doors were common. Fresh rat droppings also were found along the walls and in the storage bins inside of the barn. 33 The testing program, conducted outside of the hog barn to prevent any hazard to the hogs maintained inside the barn, was divided into three phases: 1) pre-treatment census, 2) treatment, and 3) poet-treatment census. Microencapsulated zinc phosphide 30-D bait was used for control of rats and mice. The efficacy of this rodenticide was evaluated by the percentage reduction of bait consumption between pre-treatment and post treatment censuses (ASTM, 1977b?EPA, 1977). Covered metal containers (25 x 20 x 15 cm), each with two entrances, were used for bait stations to prevent primary hazard to non-target spe cies. They were placed at 5 to 7-meter intervals along runways, next to the walls of the hog bam, under the dense vegetative cover, and near a woodpile (Fig. 3). Bait stations for the treatment were not located in the same positions of those for pre- and post-treatment censuses to pre vent prebaiting effects. Food consumption was measured daily; baits were replenished to their original weights. A torsion balance (accurate at 0.1 g) was used for weighing food and treated baits. Temperature was recorded throughout the test period using a thermograph. A. Pre-treatment Census Pre-treatment food consumption census was carried out for four days to obtain an index to the rodent population. Twelve bait stations, each filled with 200 gm EPA-standard food, were placed as indicated in Fig. 3. An extra bait station without access by rodents and filled daily with 200 g of EPA-standard food also was placed in a test area for moisture control so that adjustments of consumption data could be made. At the end of the pre-treatment baiting program all food was removed from the bait stations, and they were left empty in the test area for a three-day 34 Figure 3. Locations of bait stations in pre- and post-treatment censuses and in treatment period during a field trial for the efficacy of microencapsulated zinc phosphide 30-D at a hog farm near Waterville, Ohio. I---- 1 3-meter distance ■ locations of bait stations in pre- and post-treatment census O locations of bait stations in treatment period 35 36 lag period to counter any prebaiting effect. B. Treatment Hog food was used as the bait carrier for the zinc phosphide roden ticide, since it was a food familiar to the rats and mice in this study area. Zinc phosphide (microencapsulated formulation number 30-D) was mixed thoroughly in hog food (obtained from this hog farm) at 1% level, using a twin-shell bait mixer for about two hours. The baiting program with toxic food was carried out in the same manner as the pre-treatment census, except that the bait was exposed for ten days, and bait stations were shifted several meters from their initial locations, as demonstrated in Fig. 3, to counter any prebaiting effect. At the end of the treatment all toxic food was removed, and bait stations were washed with soap and water to remove any residual odor of zinc phosphide. A three-day lag period was allowed before the beginning of post-treatment evaluation. C. Post-treatment Census The baiting program, duplicating the pre-treatment census, both in terms of bait station locations and use of EPA-standard food, was carried out for four days. Evaluation of the Efficacy of Zinc Phosphide An evaluation of the efficacy of zinc phosphide was based on food consumption during pre-treatment and post-treatment censuses using a simple mathematical calculation as follows (EPA, unpublished): 37 Highest daily amount of food consumed during % Control« 100 - post-treatment census ______x 100 Highest daily amount of food consumed during pre-treatment census The parameter of field efficacy of a bait was required by EPA to be 70% or better for each species for which the bait was intended. However, where baits are required to control two or more species of commensal ro dents (i.e., R. norvegicus, R. rattus, and M. musculus) the tests of one of the two or three species may be as low as 60%. Additional Data for Evaluating the Efficacy of Zinc Phosphide 1, Use of tracking boards A census technique using a tracking board (Lord et al., 1970; West et al., 1972) was carried out. Each tracking board (floor tile, 7.5 cm x 15 cm) coated with marking chalk placed inside each bait stations was replaced daily throughout the baiting program (Kaukeinen, 1978). Tracks (or foot prints of rats and mice) were counted daily and indexed as follows: 0 tracks =0 I- 5 tracks =1 . 6-10 tracks = 2 II- 15 tracks =3 16-20 tracks = 4 20+ tracks =5 The index for each day was determined by summing the individual tracking board values and plotted to determine the trend of rodent activities. 2. Trapping A trapping program was carried out after the post-treatment census 38 to determine if rats and mice were still present after the baiting program. Fifteen Tomahawk live traps (for rats) were placed near or on the same locations used for bait stations in post-treatment census, and 15 Sherman live traps (for mice) were placed near locations of treatment bait sta tions. They were set for trapping animals for two days. Fifteen Victor snap traps (for rats) initially were left unset to reduce trap shyness but were,set the third day; each was placed near a Tomahawk live trap. Fif teen Museum Special snap traps (for mice) similarly were not set until the third day; each was placed near a Sherman live trap. The traps were checked when fresh bait (peanut butter) was applied each day. The cap tured animals were identified and examined for sex, weight, and general condition. 39 RESULTS AND DISCUSSION I. Bait Shyness Phenomena A. Behaviors of Rats Toward Zinc Phosphide Baits Rats sometimes avoid a poisoned diet after surviving a sublethal dose. This behavior, known as bait shyness, was defined by Rzoska (1953) as "a cautious attitude towards food (and poison bait) experienced pre viously with harmful effect." Pest control operators often have cited rats developing shyness to zinc phosphide baits and thus reducing the efficacy of this toxicant when used in field situations (Greaves,,1966; Srivastava 1968; Marsh et al., 1970; Cornwell, 1971). However, little attempt has been made to analyze this phenomenon more closely. This experiment was designed to study feeding behavior of lab and wild rats in response to zinc phosphide bait before and after exposure to a sublethal dose. Since the LD50 of zinc phohsphide for lab rats had been reported to be about 45 mg/kg, and 40 mg/kg for wild rats (Richter, 1950), attempts were made to establish a sublethal dose of zinc phosphide bait that caused an obvious physiological response in the test animals. Three groups (n= 5 or 6) of individually caged rats were used for this study. Group one was offered zinc phosphide at 0.25% in EPA-standard food for 24 hours; group two, a 0.10% bait for 24 hours; and group-3, a 0.10% bait for 20 minutes after being deprived of food for 24 hours. Three groups of wild rats (n=6) were similarly treated. Results are summarized in Table VI. Lab rats were more susceptible to zinc phosphide than wild rats, since consumption of 22.9 to 35.3 mg/kg of the toxicant resulted in 100% TABLE VI Non-choice tests on rats (R. norvegicus) with Zn3?2 baits. * Type Body Weight (g) N % Zn3?2 Toxic Bait Toxic Bait Lethal Dose (mg/kg) Suble.thal Dose Number Initial Final in EPA Exposure Time Consumption X±S.E. Range X±S.E. Range Surviv. Lab rats 280 267 5 0.25 24 hr 8.4 75.0±12.0 42.8+107.9 0 Lab rats 285 266 5 0.10 24 hr 7.3 27.6+6.1 19.4+45.7 18.6 - 1 Lab rats* 172 173' 6 0.10 20 min 5.1 30.0±2.0 22.9±35.3 0 Lab rats* 165 193 5 0.10 10 min 2.4 23.8 12.3±2.9 6.2+17.6 4 Wild rats* 278 258 6 0.10 30 min- 9.6 46.3+10.6 35.7+56.9 27.2±4.7 16.4±36.1 4 24 hr Wild rats* 310 273 5 0.10 30 min 4.9 25.3 22.7+2.2 17.2+27.9 4 *The test animals were deprived of food for 24 hours before being offered a toxic food. o 41 mortality to lab rats, whereas 35.7 to 56.9 mg/kg was required to cause death of wild rats. The lab rat that ingested only 18.6 mg/kg of the toxicant showed symptoms of sickness, as becoming inactive and walking slowly. Its face was swollen, and it exhibited piloelection. Wild rats that survived after consuming an average of 27.2 mg/kg (range 16.4 - 36.1 mg/kg) of the toxicant showed similar symptoms. Observations of normal feeding behavior of lab rats in 10-minute exposures to ground Purina chow after being deprived of food for 24 hours are summarized in Table VII. In their first exposure to ground Purina lab chow (placed in the food bowl), lab rats spent several minutes in grooming arid ejqploring the aquaria (latency period), then sniffed around the food bowl for a few seconds, and finally began to eat. Three of the lab rats ate continuously for five to nine minutes; the others left the food bowls twice during the observation period. In subsequent exposures to Zn3P2 at 0.10% in EPA-standard food, after being deprived of food for 24 hours, three lab rats approached the toxic food and immediately ate; the other two rats spent several minutes in grooming and exploring the aquaria (latency period), then approached the food bowls, and ate. The feeding behavior patterns of lab rats in 10 min e^qposures to Zn2P2 at 0.10% in EPA-standard food are summarized in Table VIII. Although the frequencies of feeding on a 0.10% bait were signifi cantly more than the frequencies of feeding on ground Purina lab. chow, the total amounts of bait ingested by lab rats in each exposure were not significantly different (Table IX). This indicated that lab rats persis ted in feeding on bait containing 0.10% zinc phosphide. TABLE VII Feeding behavior patterns of lab rats in 10-min exposures to ground Purina chow.* Frequency (one occurrence/one minute) Rat # Latency Period Feeding Interval** Feeding Interval* Feeding Total Feeding 1 1 2 2 1 2 2 5 2 2 3 1 2 1 1 6 3 5 5 - - - 5 4 1 9 - - - 9 5 4 6 6 *Animals were starved for 24 hours prior to exposures. **Interval = minute of non-feeding (grooming, exploration, etc.) Feeding behavior patterns of lab rats (same rats in Table VII) in 10-min exposures to Zn3P2 0.10% in EPA-standard food.* Frequency (one occurrence/one minute) - Rat # Latency Period Feeding Interval** Feeding Total Feeding Frequency 1 0 5 1 4 9 2*** 0 10 - - 10 3 3 2 1 4 6 4 2 .8 - - 8 5 0 10 10 ‘ J ♦Animals were starved for 24 hours prior to exposures. ■t» ♦♦Interval = minute of non-feeding (grooming, exploration, etc.) w ♦♦♦Death within 24 hours after exposure to toxic food. TABLE IX J Feeding behavior patterns of lab rats (same rats in Table VII) in 10-min exposures to Zn3P2 at in EPA-standard food.* Frequency (one occurrence/one minute) Rat # Latency Period Feeding Interval** Feeding Total Feeding Frequency 1 0 3 1 6 9 2 - - - - - 3 0 1 2 7 8 4 0 10 - - 10 5 0 10 10 *Animals were starved for 24 hours prior to exposures. **Interval = minute of non-feeding (grooming, exploration, etc.) 45 About four hours later the rats showed toxic symptoms. They became inactive and walked slowly, their faces swelled, and they exhibited pilo election. Muscular coordination was affected. One rat unsuccessfully attempted to jump up on the nesting jar; a healthy rat could do this readily (Fig. 4). Four out of the five rats survived (Table VI). Five days after surviving a sublethal dose of zinc phosphide at 0.10% in EPA-standard food these same lab rats did not reject a 1% bait in EPA-standard food. They readily ate the toxic food (Table IX); all of them died 80 to 170 minutes later (Table X). Before death the poisoned rats walked unsteadily, dragged themselves along the floor of the aquaria, lay with the belly down, extended their legs and bodies, and finally convulsed (Fig. 5). In a non-choice situation lab rats that survived a sublethal dose of zinc phosphide at 0.10% in EPA-standard food readily accepted a lethal dose in the same bait in subsequent exposure; the amounts of bait ingested by rats in each exposure were not significantly different (Table X). Thus lab rats did not show shyness to zinc phosphide baits under a non choice test situation. Observations of feeding behavior of wild rats in 30-min exposures to ground Purina lab chow after the rats were deprived of food for 24 hours are summarized in Table XI. In ejqposure to ground Purina chow wild Norway rats did not immediately eat the food when presented. They spent one to four minutes in grooming before approaching the food bowls. They sniffed about the food bowls for a few seconds, then started to eat. During 20 minutes of observation wild rats several times visited the food bowls and spent one to 14 minutes in eating for each visitation. 46 I I Figure 4. A healthy rat stands on a jar with ease. Figure 5. A lab rat that survived a sublethal dose of Zn3?2 at 0.10% in EPA-standard food exhibited muscular spasms before death after consuming Zn3?2 at 1% in EPA-standard food in subsequent exposure. 47 Figure 5 TABLE X Comparison of feeding behavior and placebo and toxic bait consumptions by lab rats in three 10-minute experiments in non-choice situations. Animal Sex Body Weight (g) Frequencies of Feeding**+ Bait Consumption (g)**++ Time to Death „ Number Initial Final I II HI I II III (After Eating ZnqP?) 1 F 168 187 5 9 9 2.5 2.5 1.3 170 min 2 F 168 165 6 10 - 1.2 4.0* 3 M 162 194 5: 6 8 1.0 1.0 1.8 83 min 4 M 158 190 9 8 10 2.7 1.8 2.3 80 min 5 M 159 228 6 10 10 3.0 2.8 2.1 125 min X±S.E. 165+2 193+10 6.2+0.7 8 .6+0.7 9.3±0.5 2.1+0.4 2.4+0.5 1.9±0 .2 114.5+21.2 ♦The rat died within 24 hours after consuming Zn3P2 at 0.10% in EPA-standard food. **I=ten-minute exposure to ground Purina lab chow (Table VII), II=ten-minute exposure to Zn3P2 at 0.1.0% in EPA-standard food (Table VIII), III=ten-minute exposure to Zn3P2 at 1% in EPA-standard food five oo days after surviving eating Zn3P2 at 0.1% in EPA-standard food (Table IX). t^-test (Ps0.05) I vs II significantly different; II vs III not significantly different t-test (P£0.05) I vs II and II vs III not significantly different TABLE XI Feeding behavior patterns of wild rats in 30-min exposures to ground Purina chow.* Frequency (one occurrence/one minute) Rat # Latency Period F I F I F I F Total Frequency of Feeding 1 1 3 1 7 5 4 5 4 18 2 1 8 5 4 3 3 6 - 15 3 4 14 5 1 10 2 4 17 4 4 5 4 1 10 2 4 - 8 5 3 3 5 4 3 5 7 - 12 F = Feeding, I = Interval (minute of non-feeding (grooming, etc.) *Animals were starved for 24 hours prior to exposures. vo 50 In subsequent exposure to zinc phosphide at 0.10% in EPA-standard food, wild rats showed feeding behavior similar to that of lab rats (Table XII). Except for animal #2 who ate almost immediately, the rats spent four to 13 minutes in grooming before eating. Rat #3 fed on the toxic food continuously for 22 minutes, while the other rats visited the food bowls several times, spending from one to seven minutes in eating the toxic food at each visitation. In a non-choice situation wild rats accepted Zn3P2 at 0.10% in EPA food as well as they had ground Purina chow; the frequencies of eating and the amounts of bait taken by rats in each exposure were not significantly different (Table XIII). The toxic symptoms, noticeable the following day, were similar to those of lab rats. However, jumping up or standing on the jar was not seen in wild rats, since they normally preferred to crouch behind or inside it. One of the test rats died (Table XIII). The four surviving wild rats were esqposed after five days to zinc phosphide at 1% in EPA-standard food; three of them survived. Their frequencies of feeding are summarized in Table XIV. Though the frequencies of eating the toxic baits by wild rats in ejqoosure to Zn3P2 baits at the two concentrations were not significantly different (Table XIII), the trend was toward fewer contacts with the 1% bait. Consumption of the 1% bait was significantly less than that of the 0.10% bait. In this study using a non-choice regimen lab rats that survived a sublethal dose of zinc phosphide bait did not reject a toxic bait in subsequent exposure; thus bait shyness was not demonstrated. Some, but not all, wild rats with prior zinc phosphide bait experience avoided the TABLE XII Feeding behavior patterns of wild rats (same rats in Table XI) in 30-min exposures to Zn3P2 at 0.10% in EPA-standard food.* Frequency (one occurrence/one minute) Rat # Latency Period F I F I F I F I F Total Feeding Frequency 1** 8 2 3 2 5 2 7 1 7 2 0 5 2 1 4 3 5 3 1. 2 14 3 8 22 ------22 4 13 2 2 1 2 1 4 1 - - 5 5 4 1 2 1 2 1 6 7 1 5 15 F = Feeding, I = Interval [minute of non-feeding (grooming, etc.)] **Death after one day of exposure *Animals were starved for 24 hours prior to exposures TABLE XIII Comparison of feeding behavior and placebo and toxic bait consumption of wild rats (Eattus norvegicus) in three 30-minute experiments in non-choice situations. Animal Sex Body Weight (g) Frequencies of Feeding** Bait Consumption (g) ** Bait Left Death (d) Number Initial Final I II III I II III Overnight (After III) 1 M 372 323 18 7 - 5.5 2.4* — — - 2 F 159 183 15 14 5 3.5 3.7 0.2 0.6 - 3 F 342 276 17 22 2 5.4 7.7 0.1 0.1 - 4 F 285 234 8 5 6 3.7 3.5 2.6 0.6 1 5 M 390 348 12 15 23 4.2 7.4 2.0 0.3 » b X±S.E. 310+42 273+29 14.0+1.8 12.6±3 .0 9.0±4.7 4.5+0.4 4.9+1.1 1.2+0.6 0.4+0.1 *The rat died within 24 hours after consuming Zn3?2 at in EPA-standard food. **I = 30-min exposure to ground Purina lab chow (Table XI) , II = 30-min exposure to Zn3p£ at 0.10% in tn EPA-standard food (Table XII), III = 30-min exposure to Zn3P2 at 1% in EPA-standard food five days # after surviving from Zn3P2 at 0.10% in EPA-standard food (Table XIV) . +t-test (P<0.05) I vs II not significantly different; II vs III not significantly different t-test (PiO.05) I vs II not significantly different; II vs III significantly different TABLE XIV Feeding behavior patterns of wild rats (same rats in Table XI) in 30-min exposures to Zn3P2 1% in EPA-standard food.* Frequency (one occurrence/one minute) Rat # Latency Period F 1 F I F I F I . F Total Feeding Frequency 1 2 0 5 25 5 3 9 1 7 1 12 - 2 4 5 1 2 2 1 1 13 2 3 - 6 5 5 2 2 4 1 8 4 9 23 F ='Feeding, I = Interval [minute of non-feeding (grooming, etc.)] m ♦Animals were starved for 24 hours prior to exposures w 54 toxic bait in a subsequent ejqxosure. However, these rats needed to con sume some toxic bait before rejecting it (Table XIII). Thus bait shyness in some of the wild rats was evidenced. Hunger might have countered the display of bait shyness, since the rats were starved for about 24 hours before the presentation of the toxic food. However, those rats with a strong aversion to the zinc phosphide bait still ingested some of the toxic food before rejecting further feeding. In other studies Prakash and Jain (1971) reported that two species of gerbils (Tatera indica and Meriones hurrianae) developed bait shyness to zinc phosphide after ingestion of a sublethal dose of bait. According to Heinz (referred to in Gaffrey, 1953), zinc phosphide was not rejected by Rattus rattus when exposure was repeated after a sublethal dose. Simi larly, Bhardwaj and Khan (1978) found that black rats (Rattus rattus) that survived sublethal doses of zinc phosphide bait did not develop aver sion to zinc phosphide rodenticides. However, Barnett et al. (1975) . reported that black rats (Rattus rattus) surviving a sublethal dose of zinc phosphide bait avoided feeding on the toxic bait on the following days. Rana et al. (1975) also suggested that the hairy-footed Gerbil (Gerbillus gleadowi) associated the sickness developed after obtaining a sublethal dose of zinc phosphide with the food mixed with the toxic compound. ’ The development of bait shyness seems to be very complicated. There are variations depending upon the nature of the bait material, the poison, and individual behavior (Rzoska, 1953). Jackson (1974) stated that rats and mice use cues, such as taste or odor, in learning to avoid poisoned food by associating their illness from a sublethal dose with that particular food or bait. 55 Although rats may use olfactory stimuli in the rejection of a toxic food, olfactory cues are not essential to forming a taste aversion un less followed immediately and repeatedly by illness (Hankins et al., 1973). Bull (1972) also indicated that odor was unimportant in the feeding beha vior of rats and suggested that initial attraction (or rejection) by odor must be reinforced by taste. Smith and Balagura (1969) stated that the taste cue was necessary for learned aversion; if the oral cavity was by-passed by stomach tube administation, conditioned aversion would not be obtained. Prakash and Jain (1971) indicated that the gerbils did not reject Zn3?2 bait in sub sequent exposure if they were administered the toxicant in sublethal-dose quantities by stomach tube initially. Nachman and Hartley (1975) reported that the effectiveness of a drug in producing a learned aversion had no relationship to the intensity of the sickness of the rats. For example, red squill produced profound behavioral disturbances (e.g., body writhing) but was not as effective as lithium chloride, which produced only mild diarrhea. Nachman and Ashe (1973) tested learned taste aversion in rats, using LiCl as an illness- producing agent, and reported that this substance.. was equally effective in producing learned aversion whether administered intraperitoneally, subcutaneously, or by stomach tube. They also indicated that the strength of the aversion was dependent upon the quantity of lithium chloride. For instance, the threshold dose for producing an aversion was about 0.15 mEq/kg, while a very strong aversion occurred at a dose of 3 mEq/kg. After being poisoned rats tended to avoid a novel food rather than a familiar food (Rozin, 1968). Kalat and Rozin (1970) found that rats formed stronger aversions to some novel solutions than to others. They 56 indicated that "salience" (the relative tendency of a novel solution to be associated with a poison) was independent of relative probability. For example, the most salient solution, casein hydrolysate, is the least preferred, while the second most salient, sucrose, is the most preferred (compared to the other three novel solutions» casein hydrolysate, NaCl, and vanilla solutions). They concluded that the salience of a solution might depend on three factors: specific chemical properties, strength of stimulation, and a special case of novelty. Kalat (1974) reported that the association by rats of a given taste with a toxicant depended largely on the novelty of the taste not on the "strength" of the taste. Kalat and Rozin (1973) proposed a concept of "learned safety," that is, that rats learned to accept a food that caused no ill effects. They tested groups of rats with LiCl by intraperitoneal injections at various delay times after the rats had drunk a casein hydrolysate solution and reported that the rats poisoned 1/2 hour after drinking a solution had stronger aversion to the solution than those that were poisoned four hours or more after drinking. They indicated that the rats were learning during the delay that the ingested solution was "safe." Nachman and Jones (1974) confirmed this concept with similar experiments. Learned taste aversion or bait shyness was classified as a classical conditioning by Seligman (1970) in that rats were prepared to associate taste with illness, even over long delays of reinforcement, but were oontraprepared to associate taste with footshock. Besides the development of bait shyness, some wild rats may possess neophobic behavior, which protects them from eating poisoned diet in their natural surroundings (Barnett, 1958); Chitty and Shorten (1946) first 57 described such behavior of rats as an avoidance of unfamiliar objects in familiar surroundings. They also stated that only wild rats, not domestic rats, exhibited such avoidance behavior. Social behaviors may have influence on poison avoidance of the young rats. Galef and Clark (1971) demonstrated that adult rats would "lead” their offspring to feed on a "safe" diet in an environment containing a poisoned diet that the adults had had experience with. They also indica ted that the young rats appeared to use olfactory and visual cues combined with neophobic behavior in avoiding the toxic food and also continued to avoid that diet after they were isolated frofti the parent in a new environment. The present study indicated that in a non-choice situation lab rats did not develop shyness to zinc phosphide bait. However, some wild rats in a similar situation avoided zinc phosphide bait in subsequent exposure. The animals appeared to use taste cues in associating their illness with the zinc phosphide bait, since they did not reject the poisoned food on subsequent exposure after sniffing it but showed aversion to zinc phosphide bait only after consuming some of the toxic bait. B. Nature of Bait Shyness In earlier experiments (Table XIII) some (though not all) wild Norway rats which survived a sublethal dose of zinc phosphide bait rejected such toxic bait in subsequent exposures. However, what actual components (zinc phosphide, EPA standard food, or even both) the rats responded to are still in question. Three groups (n=4-6) of wild rats which had survived a sublethal dose 58 of zinc phosphide at 0.10% in EPA standard food were used for determining types of bait shyness. Group I was retested with zinc phosphide at 1% in EPA standard food; group II, with Zn3P2 at 1% in Bait X; and group III, with EPA standard food, in a non-choice situation. The results are sum marized in Table XV. Six wild rats in group I that survived a sublethal dose of zinc phos phide at 0.10% in EPA standard food consumed an average of 2.2 g of Zn3P2 at 1% in EPA on subsequent exposure. This was significantly less than their average daily consumption of ground Purina chow on the fifth day after surviving a sublethal dose, and five of them survived. Five surviving rats in group II consumed an average of 1.4 g of Zn3P2 at 1% in Bait X on sub sequent exposure. This also was significantly less than their previous average daily consumption of ground Purina chow, and four of them survived. Thus, these wild rats developed shyness specifically to zinc phosphide, since they rejected this compound mixed with an alternate bait material. Wild rats in group III that survived a sublethal dose of zinc phos phide at 0.10% in EPA standard food accepted EPA standard food without toxicant on subsequent exposure, consuming an average of 12.1 g. Although this consumption of EPA standard food on subsequent exposure was signifi cantly less than the previous daily consumption of ground Purina chow, it was much greater than the consumption of toxic baits by the other two groups. In this study wild rats in non-choice situations developed specific shyness to the rodenticide but not to the bait material. In other studies, lab rats previously experienced with barium car bonate and/or arsenic baits did not reject those rodenticides when offered in a new bait base (Rzoska, 1953). Consequently, he indicated that the food medium was responsible for a subsequent reluctance or refusal to eat TABLE XV Tests of shyness to wild rats (Rattus norvegicus) in a non-choice situation to different kinds of baits Group Body Weight (g) Mean Consumption (mg/kg) Number One Day Consumption (X±SE) Mean mg/kg Number (N=6) Initial Final Exposure to Zn3P2 at .1%* Survivors GPC** Zn3P2+ Sn3P2+ EPA Sub- Leth. Survivors Sublethal Lethal In EPA in X*** lethal I 328 328 21.7 - 6 26.5 vs 2.2++ 68.3 59.8 5 ±1.0 ±0.2 II 280 277 23.2 14.0 5 26.0 vs 1.4 46.2 42.4 4 ±3.0 ±0.5++ - , _ _ ++ Ill 273 254 21.4 35.5 4 21.9 vs 12.1 4 ±2.0 ±3.6 *In EPA, 30-minute to 24-hour exposure **GPC - Ground Purina chow on the fifth day of survivi ***X = 94% ground barley and 5% vegetable oil, by weight 5 +Zinc phosphide concentrations at 1% ++Significantly different (t-test, P<0.05) 9 60 (Bhardwaj and Khan, 1978). Prakash and Ojha (1977) reported that the Indian gerbil (Tatera indica) developed aversion only to zinc phosphide rodenticide. Their using different bait materials with zinc phosphide did not reduce the occurrence of bait shyness. In contrast, another toxicant (RH-787) used with the same bait was readily consumed by this species in a second exposure. Howard et al. (1977) stated that deer mice (Peromyscus maniculatus), which developed bait-shyness during the first control program, might retain their aversion to the toxic bait or only the bait (i.e., oats) for the rest of their lives. However, they indicated that prebaiting with clean oats could condition some 1080-shy rodents to feed again on a bait they had been conditioned to avoid through eating a sublethal amount. C. Retention of Memory for Bait Shyness The interval that wild Norway rats experienced with zinc phosphide bait can retain memory of this toxic food is not well studied, and an experiment was designed to evaluate this phenomenon more closely. The results are summarized in Table XVI. Fourteen days after surviving a sublethal dose of Zn3P2 at 0.10% in EPA standard food, all rats in group I consumed an average of 1.3 g # of 1% zinc phosphide in EPA standard food and survived. At 30 days two out of four rats similarly treated survived; at 60 days, two out of three survived. Thus the memory retention for zinc phosphide bait by some wild Norway rats lasted for at least 60 days. Although there was a trend of higher toxic bait consumption on the subsequent exposure of rats in group III Edue to heavy consumption (6.7 g) by one rat who finally died] than in the other groups, there was no significant difference in 60 (Bhardwaj and Khan, 1978). Prakash and Ojha (1977) reported that the Indian gerbil (Tatera indica) developed aversion only to zinc phosphide rodenticide. Their using different bait materials with zinc phosphide did not reduce the occurrence of bait shyness. In contrast, another toxicant (RH-787) used with the same bait was readily consumed by this species in a second exposure. Howard et al. (1977) stated that deer mice (Peromyscus manieulatus), which developed bait-shyness during the first control program, might retain their aversion to the toxic bait or only the bait (i.e., oats) for the rest of their lives. However, they indicated that prebaiting with clean oats could condition some 1080-shy rodents to feed again on a bait they had been conditioned to avoid through eating a sublethal amount. C. Retention of Memory for Bait Shyness The interval that wild Norway rats experienced with zinc phosphide bait can retain memory of this toxic food is not well studied, and an ejqperiment was designed to evaluate this phenomenon more closely. The results are summarized in Table XVI. Fourteen days after surviving a sublethal dose of Zn3P2 at 0.10% in EPA standard food, all rats in group I consumed an average of 1.3 g # of 1% zinc phosphide in EPA standard food and survived. At 30 days two out of four rats similarly treated survived; at 60 days, two out of three survived. Thus the memory retention for zinc phosphide bait by some wild Norway rats lasted for at least 60 days. Although there was a trend of higher toxic bait consumption on the subsequent exposure of rats in group III Eclue to heavy consumption (6.7 g) by one rat who finally died] than in the other groups, there was no significant difference in TABLE XVI Retention of memory for bait shyness of wild rats (Rattus norvegicus) to zinc phosphide baits after surviving a sublethal dose Group Body Weight (g) Number Zn3P2 at 0.10% Number Days before Gonsunption Number Initial Final Animals in EPA Exposure Survivors Reexposure to Z113P2 (1% in EPA) Surviv. Sublethal4" Lethal4" Zn3P2(l% in EPA) X±SE Sub.4" Lethal1" I 278 258 6 27.2 46.3 4 14 1.3±0.6* 60.2** 4 II 261 259 5 16.7 14.3 4 3° 1.6 ±0.7* 36.1 69.9 2 III 280 264 6 22.3 40.8 3 60 2.5+2.1* 15.2 394.1 2 Exposure for 30 min- 1 day *No significant difference in toxic bait consumption between groups I, II, and III (ANOVA, 95% con fidential limit) **0ne animal in this group consumed 152 mg/kg of toxicant and inexplicably survived. *4“ Mean mg/kg 62 toxic bait consumption between these three groups. According to the present study it can be concluded that wild Norway rats, but not lab rats, after obtaining sublethal amounts developed shy4- ness to zinc phosphide bait (particularly to the rodenticide and not the bait material), and some retained memory for the toxic bait for at least 60 days. Some wild Norway rats may retain memory for zinc phosphide bait for longer periods; therefore, use of larger sample sizes and the extension of the interval between exposures to zinc phosphide baits are suggested for further investigation. Memory retention by other rodents after exposure to zinc phosphide has been studied by Prakash and Jain (1971). They reported that the hairy-footed gerbil (Gerbillus gleadowi) retained memory for zinc phosphide bait for only 10 to 15 days. Mohan Rao and Rajabai (1978) reported that the Indian field mouse (Mus booduga) and the spiny field mouse (Mus platg- trix) developed bait shyness to zinc phosphide after one day’s exposure to a sublethal dose; the aversion persisted for 95 days in the former and 75 days in the latter. In other studies, Rzoska (1953) reported that lab rats retained bait shyness for red squill for 2 to 53 weeks, but with great individual varia tions; for arsenic, over a month; and for barium carbonate, two months. Howard et al. (1977) found that deer mice (P. maniculatus) retained memory for compound 1080 bait for eight months, although the shyness to this toxic bait decreased with time. D. Social Behavior Towards Bait Shyness The purpose of this study was to determine feeding behavior of wild 63 Norway rats in a social group in response to non-toxic bait (ground Purina chow) and zinc phosphide bait after surviving a sublethal dose of this toxic bait. Seven wild rats were used for this study and kept in a test room (Fig IB). Animals were provided with ground Purina chow at night only (the start of the red-light cycle) on the six days before the initiation of observations. After release into the test room, one rat died on the second day, probably from fighting with other rats, since blood was found on the wall and on the wooden boxes; its head had wounds. At time of bait presentation the occurrence of feeding behavior of each rat was observed and recorded in each minute during a 60-minute observation period. Other behaviors (e.g., grooming, exploring the sur roundings, fighting, and chasing) that occurred also were observed and recorded. The patterns of feeding behavior of these wild rats during the first observation period are summarized in Table XVII. The rats did not imme diately approach the ground Purina chow at the time of food presentation; they groomed and explored the nest boxes and the test room. On the eighth minute the dominant male (rat #4) approached the food source; it sniffed the food for a few seconds and then started to eat. Four minutes later another male (#1) and a female (#6) approached the food source and ate together with the dominant male. One minute later the other two males (#3 and #5) joined the group and fed. The dominant male rat ate continuously for 21 minutes, then paused for 13 minutes, and returned to eat for one more minute; then he stopped eating the food again for 12 minutes before returning to eat continuously for about one to eight minutes, then paused for one to 24 minutes before TABLE XVII Pattern of feeding behavior of a colony (n=6) of wild rats in 60-minute exposure to ground Purina chow. Rat # Sex Latency Period FIFIFI FIFIFIFIF Total Feeding Freq. (minutes) ' : . 1 M 12 5 24 7 7 5 17 2 M 60 3 M 13 8 3 6 13 3 1 1 5 2 4 1 21 4 M 8 21 13 1 12 5 27 5 M 13 5 1 1 1122 15 1517113 15 6 F 12 3 16 3 1 3 1 2 19 11 F = feeding, I = Interval [non-feeding time, (grooming, etc.)I 65 returning to eat again. One male rat of this social group did not approach the food source at all; it sat quietly in one spot of the test room during the first observation. These rats were transferred to individual cages and fed with Zn3P2 at 0.1% in EPA standard food for 30 minutes to one day. Three of them died one or two days later; one of these was the dominant rat. Five days after consuming a sublethal dose of zinc phosphide bait (an average of 22.9 mg/kg) the three survivors (one female and two males) were returned to the test room. On the following day they were exposed to Zn3P2 at 1.0% in EPA standard food in a non-choice situation. The patterns of feeding behavior of these three survivors in response to a 1% bait are summarized in Table XVlu. The female rat first approached the toxic bait at the fifth minute after the toxic bait was presented, investigated, and sniffed it for a moment, then further ignored it. However, 25 minutes later she returned to the toxic bait, tasted it once, and then again rejected it. The new dominant male (rat #3), which weighed about 364 g, approached the toxic bait on the eighteenth minute of exposure, sniffed for three to five seconds, and started eating. It ate the poisoned food continuously for two minutes, then paused for three minutes before returning to the toxic food again. This male rat visited the toxic food several times and ate for one to three minutes at each visitation. The subordinate male (rat #1), which weighed about 222 g, tried to approach the bait station several times but was chased away by the dominant male. The dominant male spent about 11 minutes eating the poisoned bait alone and ingested a total TABLE XVIII Pattern of feeding behavior of a colony (n=3) of wild rats that survived a sublethal dose of Zn3P2 at 0.10% in EPA for 6 days in ejq?osure to Zn3P2 at 1% in EPA in 60-min observations. Frequency (one occurrence/one minute) Rat # Sex Latency Period F I F I F I F I F I F I Total Frequency (minutes) of Feeding 1 M 60 0 3 M 18 2 3 2 1 1 4 2 2 3 4 1 17 11 6 F 30 1 29 1 F = feeding, I = interval Enon-feeding time (grooming, etc.)3 »j 68 of about three grains during the 60-minute observation. By the next morning a total of 13.2 g had been eaten; both of the male rats were dead, and only the female survived. According to the toxic bait consumption, it might be hypothesized that 1) the dominant male returned to eat more toxic bait, 2) the subordinate male probably waited until the dominant male became sick or died, then approached the toxic bait and consumed it without hesitation, 3) the female may have returned to eat some toxic bait but did not take enough to be killed. During the observation period the female tasted the toxic food only one time and then rejected it. She did not come back to visit the toxic bait again and never tried to chase the dominant male away during the several times he returned to feed. This suggested that the development of this bait shyness was an individual behavior, not influenced by the actions of other animals. However, in other studies Galef and Clark (1971) reported that the offspring born in a colony and with no experience with poisoned food learned to avoid poisoned food by following their parent to feeding sites and eating the food selected by the adults. In a larger colony it is possible that inexperienced young animals may follow a dominant male, who has had experience with toxicant baits, to feeding sites and avoid eating toxic foods merely by imitating the dominant male in rejecting a food. In a stable population, a dominant male> who has had experience with poisoned food and who nests near the feeding sites, associates with several females and their offspring and excludes subordinate males (Calhoun, 1962), may influence bait shyness development in his social group by being followed by females and their offspring to feeding sites where the food is safe. Subordinate males are 69 more likely to range widely and be exposed to toxic baits. Further investigation is needed to understand these phenomena. According to this study lab rats did not reject zinc phosphide bait on subsequent exposure in a non-choice situation. However, some wild rats that survived a sublethal dose of zinc phosphide bait avoided the zinc phosphide bait on subsequent exposure; and some wild rats retained memory for zinc phosphide for at least 60 days. Pest control operators often have cited rats developing shyness to zinc phosphide baits as reducing the efficacy of zinc phosphide used in field situations (Greaves, 1966; Srivastava, 1968; Marsh et al., 1970; Cornwell, 1971). Therefore, if the rat control programs using zinc phosphide bait were found to be unsuccessful, a second bait distribution should not be made within two months. Merely changing the bait material . ■ ■ for the second operation likely will not be successful, since the bait-shy rats largely developed aversion to the zinc phosphide rodenticide and not to the bait material itself, II. Improving Toxic Baits to Increase the Efficacy of Zinc Phosphide A. Bait Developments Several kinds of cereals, such as wheat (Srivastava, 1968), oat groats (Clark, 1975), barley (Dana, 1962), cracked com (Rudd and Genelly, 1956), and polished rice (West et al., 1972) have been used as bait carriers for zinc phosphide in rodent campaigns. However, in some cases such baits did not achieve control of rodents; the reduced efficacy might have resulted from poor initial bait acceptance (West et al., 1972). To overcome this problem, attempts have been made to improve the 70 palatability (taste or odor that is accepted) by adding substances, such as oils and sugar, so that the finished baits are more readily accepted by rodents. The present EPA-standard food, consisting of 65% cornmeal, 25% oat groats, 5% corn oil, and 5% powdered sugar, is palatable to rats and mice. While it is used as a challenge diet in choice tests for determining the efficacy of rodenticides under laboratory conditions, it is not the most accepted diet by rats and mice. Therefore,,more research is needed on developing preferred bait formulations. In the present study two kinds of bait formulations were tested on lab and wild rats. One involved the addition of enhancers (substances that act as appetizers), such as salt, red pepper, sugar, at different concentrations to EPA-standard food; the other, the use of different bait materials (e.g., ground corn, barley, wheat, and soybeans) in different combinations. The results of testing lab rats with EPA-standard food and EPA-standard food combined with enhancers are summarized in Table XIX. Red pepper at 0.16% and coconut flakes at 1.25%, 2.5%, and 5% increased palatability of EPA-standard food for lab rats. Granulated sugar and brown sugar at 10% levels in EPA (which lacked the normal pow dered sugar) also increased palatability significantly. However, both substances did not prove to be better than the powdered sugar formulation of the EPA standard food when they were mixed at 5% in EPA (which lacked the normal sugar). Lab rats did not show preference between EPA-standard food and EPA (which lacked the normal powdered sugar) containing 2.5% brown sugar. EPA (lacking powdered sugar) combined with 2.5% coconut flakes and 2.5% brown sugar also was not preferred to EPA-standard food by lab rats. Thus, the texture of sweetness enhancers had little influence 71 TABLE XIX Consumptions of EPA standard baits modified by addition of enhancers by addition of enhancers by lab rats in choice tests, five ani mals per test group. 4-Day Consumption (g) (X+SE) Enhancers % in EPA EPA* EPA*+Test t-test Substance (P<0.05) 1. Salt 0.16 40.6+2.0 35.0+3.3 NS 2. Salt 0.31 28.9±6.1 26.3+5.6 NS 3. Salt 0.63 34.8±2.3 29.1±5.2 NS 4. Salt 1.25 31.8±7.1 21.9+3.9 NS 5. Salt 2.5 39.8+8.1 14.7±6.8 + 6. Salt 5.0 47.6+9.8 7.5±2.3 + 7. Red pepper 0.16 32.1±3.6 46.4+4.7 .+ 8. Red pepper 0.31 36.3+7.4 25.7±6.9 NS 9. Red pepper 0.63 33.6±4.1 27.6+5.2 NS 10. Red pepper 1.25 40.9+1.5 20.7±2.0 + 11. Red pepper 2.5 37.8±6.9 7.9±3.0 + 12. Red pepper 5.0 48.0±8.7 6.0+1.4 ' + 13. Coconut flakes 1.25 24.7±1.3 30.9±1.6 + 14. Coconut flakes 2.5 14.3±3.0 42.1±2.0 + 15. Coconut flakes 5.0 18.0±2.9 47.1Ü.1 + 16. Granulated sugar 5.0 31.9+6.0 33.6±5.5** NS 17. Granulated sugar 10.0 11.5±2.8 46.9±8.9** + 18. Brown sugar 2.5 34.6±3.1 38.6±7.1** NS 19. Brown sugar 5.0 23.5±3.3 35.2±6.5** NS 20. Brown sugar 10.0 14.0±1.4 45.4+2.9** + 21. Coconut flakes 5.0 29.1+2.5 35.7±6.3** NS 22. Coconut flakes+brown sugar 2.5+2.5 35.7±3.4 37.9+5.8** NS 23. Monosodium glutamate 1.0 37.5±4.7 19.8±4.0*** + 24. Garlic powder 1.0 40.8+5.2 12.3+1.5*** ■ + Difference in consumption significantly different NS = Difference in consumption not significantly different ♦Consists of 65% ground corn, 25% rolled oats, 5% com oil, and 5% powdered sugar **EPA lacked powdered sugar ***EPA consists of 65% ground com, 25% rolled oats, 5% vegetable oil, and 5% powdered sugar 72 on a choice feeding of lab rats, nor was the 2.5% concentration important. Salt did not increase palatability of EPA standard food in all cases. Monosodium glutamate and powdered garlic at 1% in EPA were rejected by lab rats. Red pepper probably acted as an appetizer by stimulating salivary gland and gastrointestinal activities; however, when it was mixed at higher concentrations, it reduced food consumption. Perhaps its pungent taste was too strong for the rats. In other studies, Hilker et al. (1967) tested lab rats with spiced diets (containing spices, such as ginger, cloves, cinnamon, and black pepper) and an unspiced diet. In a choice situation they found that adult rats did not show preference for any of the individual spices (ginger, cloves, cinnamon, each at 5% by weight; or black pepper, cloves, cinnamon, each at 0.5% by weight), nor did they select (or reject) foods with combined spices. However, young rats preferred unspiced diets. Schein and Orgain (1953) also reported that rats avoided foods that were highly spiced. Coconut flakes added to EPA-standard food at any concentation consistently increased palatability for lab rats, probably because of the sweet taste. In other studies, Crabb and Emik (1946) reported that ground fresh coconut mixed with rolled oats at ratios of 1:4 to 1:6 by weight was well accepted by both Polynesian rats (Rattus exulans) and black rats (R. rattus). Shuyler (1954) also reported that dried coconut was one of the most acceptable fruits, probably being more acceptable than cantaloupe and banana for wild Norway rats. Doty (1945) found that half-inch cubes of fresh coconut coated with zinc phosphide and mill lime (calcium hydroxide) at the rate of 0.33% of the former and 0.25% to 0.33% of the.latter were very acceptable and effective in rat campaigns in sugar 73 cane fields in Hawaii. West et al. (1972) also used l-cm^ chunks of fresh coconut dusted with zinc phosphide (1% by weight) for rat control in the Philippines. Strecker et al. (1962), in testing bait bases for use with poisons in controlling Polynesian and roof rats on Ponape, E. Caroline islands, found that under choice conditions in the fieid toasted coconut was preferred to dry rice, or ground number 3 copra (the poorest grade). Salt was a substance that provided negative reactions in most cases. While not totally rejected by lab rats at lower concentrations, palata bility was not increased. A salty taste was not necessary for the rats to increase bait consumption. In other studies, however, salt solutions (7.9 g/L sodium chloride) were found preferred to tap water by rats (Garcia et al., 1971). Richter (1939) reported that normal rats detected salt in concentrations as low as 0.055% and drank more of these solutions than distilled water. Young (1932) maintained test rats for his study with the diet containing salt at 0.5% by weight. Shumake et al. (1971) reported that both lab and wild rats showed progressive aversion to wheat pellets treated with sodium chloride in direct proportion to the increase of salt concentrations. Monosodium glutamate at 1% also was found to reduce consumption. This compound was introduced into United States in 1948 from Japan in foods for human consumption, as it enhanced food acceptability (Melnick, 1950). He indicated that this compound, with a meatlike flavor, had four taste components (sweetness, sourness, saltiness, and bitterness); it not only increased salivary secretion but also stimulated a tingling feeling in the mouth, and the total taste sensations persisted. Studies by Scott and Quint (1946) on lab rats, given a choice between a standard diet, showed neither like nor dislike by rats for this flavor. If such 74 results can be generalized, then the vegetable oil component of the bait might have been a factor in reducing bait consumption; however, no compari son was made of lab rat preferences between vegetable and corn oils. Garlic powder at 1% was another substance that resulted in reduced consumption, perhaps because of its pungent taste. In contrast to this study, Oderkirk (1961) reported that garlic powder, when used in very small (but unspecified) amounts, improved acceptance of red squill bait for wild rats by masking the squill’s bitter taste. Granulated sugar and brown sugar at 10% increased palatability sig nificantly. This agreed with the results of Shumake (1978) who found that sweetness enhancers (e.g., sugar) increased the bait consumption of the rice-field rats (R. r. mindanensis). In other studies, Hausman (1933) reported that lab rats preferred sugar solutions (20% to 40% sugar in solutions) to pure water. Richter and Campbell (1940), testing taste preferences of lab rats for five common sugars, found that rats showed the greatest preference for maltose, with maximum concentration in the solution at 11%, next for glucose at 10% and sucrose at 8%, and only slight preference for galactose at 9%, but none for lactose. Young (1932) reported that, given a choice, rats preferred sugar to butter-fat, wheat, or flour. Shumake et al. (1971) reported that both lab and wild rats showed increasing preferences for wheat pellets treated with sucrose in direct proportions to sucrose concentrations. Those bait enhancers that increased palatability of EPA-standard food for lab rats were tested on wild rats. The results are summarized in Table XX. Red pepper at 0.16% increased palatability of EPA-standard food for wild rats, but coconut flakes at 2.5% and 5%, granulated sugar at 10%, and brown sugar at 10% did not. TABLE XX Average 4-day food consumption of wild rats (R. norvegicus) in choice tests using experimental baits and EPA-standard baits, and EPA + enhancers and EPA standard baits, n=5. Experimental Enhancer % by Weight 4-Day_Consumption (g) t-test Baits* (X ± S.E.) (PS0.05) Experimental* EPA+enhancer EPA A 26.8±10.1 28.6+5.7 NS B 47.1±7.3 46.8+10.0 NS E 50.1±10.4 40.1±13.7 NS Coconut flakes 2.5 54.4+15.6 39.1+9.4 NS Coconut flakes 5.0 47.3±12.5 37.0±12.0 NS Red pepper 0.16 56.9+9.2 34.4 ±5.7 ' + Granulated sugar 10.0 34.6+11.0** 35.4+12.1 NS Brown sugar 10.0 33.2±7.1** 41.2+10.2 NS «4 W *See Table XXI ** EPA lacked powdered sugar "^difference in consumption significantly different NS = difference in consumption not significant 76 Several bait combinations were compared on lab rats with EPA-standard food. The results are summarized in Table XXI. Baits with ground barley, a component of Bait A, and ground wheat (Bait B) had increased consumption. Those bait combinations (Bait A, Bait B, and Bait E) preferred over EPA- standard food by lab rats also were tested on wild rats. The results are summarized in Table XXI. Wild rats did not prefer any of these bait com binations over EPA-standard food. In other studies, Oderkirk (1961) tested wild Norway rats for their food preference and found that the rats preferred the corn meal-oats mix ture (ratio 50:50 by weight) to a food bait containing only corn meal or only oats. Cornwell and Bull (1967) studied food preferences of wild rats and reported that rats preferred grains in the following order: maize, wheat, and rice. They also indicated that a mixture of 50/50 of oat and maize meal was as acceptable as oatmeal alone. Khan (1974) studied the food preferences of the black rats (Rattus rattus) and reported that of cereal grains the rats showed most preference for millet, next for wheat, and then maize; ground cereals were preferred to whole cereal by rats. Shuyler (1954) reported that rolled oats and ground whole corn were the most acceptable of the 32 grains and seed materials evaluated in his study of wild rats. Crab and Emick (1946) studied the food preference of the Polynesian rat (Rattus exulans) and the black rat (Rattus rattus) in field situations on Guam and reported that rolled oats appeared to be the cereal of choice (conpared to wheat). The texture of food may have some influence upon the selection of food by rats. Young (1932) studied the food preferences of the white rats in a free choice between two foods and reported that wheat seeds TABLE XXI Bait combinations tested on lab rats in choice feeding with EPA-standard food (N=5). Exper imental Grain Combinations % by 4-Day Consumption (g) t-test Bait* Weight (X ± S.E.) (P.S0.05) Experimental EPA A ground barley + rolled oats 65+25 46.8 + 4.0 11.5 ± 3.0 + B ground wheat + rolled oats 65+25 55.2 + 2.6 22.0 + 4.7 + C ground soybean + rolled oats 65+25 12.9 ± 4.9 58.8 + 11.7 + D ground barley + ground wheat + 30+30 35,4 + 5,0 23,7 + 3.9 NS ground soybean +30 / E ground barley + ground corn + 35+35 34.5 + 2.9 17.8 + 0.7 + rolled, oats +20 F ground wheat + ground corn + 35+35 26,1 + 7.0 34.8 + 2.7 NS rolled oats +20 G ground soybean + ground corn + 35+35 18.9 + 5.5 52.2 + 6.9 + rolled oats +20 H ground barley + ground wheat + 25+25 40.8 + 3.7 34.0 + 9.2 NS ground corn + rolled oats +25+25 «4 «4 *includes 5% powdered sugar and 5% corn oil by weight difference in consumption significantly different NS = difference in consumption not significantly different 78 were preferred to flour. Barnett (1956) also reported that wild rats tended to prefer whole grains to meal. In conclusion, bait enhancers, such as red pepper at 0.16%, coconut flakes at 1.25, 2.5, and 5% increased palatability of EPA-standard food for lab rats. EPA-standard food (which lacked the normal powdered sugar) combined with granulated sugar or brown sugar at 10% was preferred to EPA-standard food by lab rats. However, similar results were not obtained with wild rats, except for red pepper at 0.16%, which was found to be a consistent attractant with wild rats. Red pepper concentrations ranging from 0.32 to 5%, salt at 2.5 to 5%, monosodium glutamate at 1%, and garlic powder at 1% reduced the bait consumption of lab rats. Bait combinations A, B, and E were preferred to EPA-standard food by lab rats, but none of them provided similar results for wild rats. Food preferences of wild rats appeared to have little correlation with those of lab rats; however, further investigation on improving the palatability of the toxic baits for rodent Control are needed. If the bait materials are more palatable to the rodents, more bait will be eaten; and the chance for bait shyness development by rodents thus is reduced. Since red pepper at 0.16% proved to increase palatability of EPA- standard food both for lab and wild rats when added to EPA-standard food, it was selected for testing, as an enhancer for atoxic bait (#2-14) on wild rats in a choice test. However, this formulation provided only 40% mortality to the test rats (Table XXII), while an unmodified zinc phosphide bait (#2-14) gave 80% mortality to the test animals (Table XXIII). Perhaps the rats associated their sickness with a pungent taste of red pepper and rejected the toxic food before consuming the lethal dose. TABLE XXII Choice tests on wild rats (Rattus norvegicus) with Zn3P2 formulations at 1% in EPA-standard food and EPA placebo bait, n=5. Zn3P£ For- Body Weight (g) 3-Day Consumption (g) t-test Mean Dose (mg/kg) Mortality Mean Time to mulation Initial Final (X 1 S.E.) # Deaths/ Death EPA Toxic Bait Sublethal Lethal Test Animal (days) 30-B# 305 300 38.6114.1 2.310.2 + 77.1 83.2 2/5 3 25-B# 290 308 36.7+11.0 2.410.5 + 67.1 117.4 2/5 3 30-D# 332 322 27.3±11.6 2.810.9 + 28.8 97.8 4/5 3 18-D# 350 357 12.719.2 2.110.4 NS 36.0 74.4 3/5 1 Zn3P2+NaHCO3 ** 317 329 18.1111.5 1.010.4 + .6.2 29.3 2/5 1 Zn3P2 2-14 256 254 12.8110.6 2.310.4 NS 94.8 92.8 2/5 ■ 1 +red pepper at 0.16% 30-D*ff 294 283 22.019.6 3.610.4 NS 45.0 74.7 3/5 2 *at 0.5% concentration **washed; at 0.5% concentration +significantly different (P<0.05) # NS = not significantly different encapsulated TABLE XXIII Choice tests on wild rats (Rattus norvegicus) and wild house mice (Mus musculus) with Zn3P2 (with different toxicant formulations) at 1% in EPA-standard food and EPA placebo baits, n=5. Type Zn3P2 # Body Weight (g) 3-Day Consumption (g) t-test Mean Dose (mg/kg) Mortality Mean Time Formulation Initial Final (X + S .E.) # Deaths/ to Death EPA Toxic food Sublethal Lethal Test Animal (days)- Wild rats 1-21 248 261 34.1±8.8 2.3+0.3 + 99.4 134.5 1/5 1 Wild rats 2-14 284 288 4.3+1.6 2.8+0.5 NS 21.0 128.7 4/5 1 Wild rats 7-11 302 299 10.0±7.2 2.2+0.8 NS 42.5 108.5 2/5 1 Wild rats 7-14 305 311 24.6+8.6 1.7+0.4 + - 54.9 5/5 1 Wild rats 4-7 298 322 61.3+15.5 1.010.3 + 19.1 95.7 1/5 1 Wild mice 2-14 21 21 6.2+2.4 0.1+0.1 + 43.1 210.5 1/5 1 o 8 0 + significantly different (P£0.05) NS=not significantly different 81 Thus a bait enhancer that increased food palatability might have a nega tive result when mixed with the toxic bait due to the distinctive taste imparted to the bait. B. Using Buffering Substances The decomposition of Zn3P2 in the rat's stomach by hydrochloric acid has been assumed (Elmore and Roth, 1943). If such an acid condition could be degraded by adding buffering substances to the zinc phosphide, the early decomposition of this compound might be slowed. The rats thus could consume more toxic bait and be killed before they associated illness with the toxic food. NaHCO3, MgCO3, and CaHPO3 were used as buffering substances and com bined with zinc phosphide for both choice and non-choice feeding on lab rats. The results are summarized in Tables XXIV and XXV. in both non-choice and choice tests using ground Purina chow as a challenge diet, all animals died on the first day. However, the use of ground Purina chow as a challenge diet raised a question. Of the two diets used as a bait carrier, the EPA-standard food was the more palatable; this may have stimulated the test rats to eat more toxic food. Therefore, zinc phosphide combined with NaHCO3 (both washed and unwashed formulations) were tested on lab rats using EPA-standard food as both the toxic carrier and a challenge diet. The results are summarized in Table XXVI. These formulations gave only 80% mortality, similar to that of the other zinc phosphide baits (both washed and unwashed formulations); thus, buffering substances used had no significant influence on the consumption of the toxic food. It was possible that the amounts of buffering ingested by the TABLE XXIV Non-choice tests using lab rats (Rattus norvegicus) eiqoosed to EPA-standard food containing 1% experimental zinc phosphide combined with buffering substances Buffering Wash Mean Animal Body One-day Consump One-day Consump Mean Dose Mortality Time to Substances Weight (g) tion of GPC* (g) tion of Toxic Bait (mg/kg) # Deaths/ Death Initial Final (X ± S.E.) (9) (X + S.E.) Test Animal (days) MgCO3 + 193 206 22.1 + 2.3 4.1 ± 0.6 213.0 5/5 1 NaHCO3 + 190 211 23.1 ± 1.0 7.2 ± 0.8 373.0 5/5 1 CaHPO3 + 191 210 24.3 ± 1.8 4.5 + 0.8 232.8 5/5 1 MgCO3 - 176 200 22.3 ± 1.1 6.7 ± 0.8 392.4 5/5 1 NaHCO3 - 187 206 22.6 ± 0.9 5.1 + 0.6 277.6 5/5 1 CaHPO3 - 188 209 24.6 ± 1.5 5.9 ± 0.6 317.1 5/5 1 - + 228 247 21.9 ± 2.3 4.4 ± 1.5 182.0 5/5 1 - — 228 225 16.2 + 2.3 3.6 ± 0.9 160.3 5/5. 1 00 K) ♦ground Purina chow TABLE XXV Choice tests using groups of lab rats (n=5) exposed to EPA baits containing 1% experimental zinc phosphide formulations and ground Purina chow. Buffering Wash Mean Animal Body 3-Day Consumption Mean dose Mortality Mean Time to Death Substances Weight (g) (g) (X ± S.E.) t-test mg/kg # Deaths/ (days) Initial Final GPC*„ Toxic Bait Test Animal MgCO3 + 166 195 7.6+3.1 2.1+0.6 NS 127.8 5/5 1 NaHCO3 + 166 197 0.2±0.1 3.3+0.7 + 200.3 5/5 1 CaHCO3 + 166 192 5.0+5.0 3.0±0.4 NS 173.3 5/5 1 MgCO3 171 198 1.6+1.3 3.9±0.7 NS 228.3 5/5 1 NaHCO3 - 168 175 7.5±5.6 2.8+0.3 NS . 169.2 5/5 1 CaHCO3 - 171 174 2.1±1.2 3.4+0.7 NS 196.9 5/5 1 - + 169 156 l.l±0.5 2.0+0.2** NS 186.8 5/5 1 - - 164 167 6.6+4.9 2.0+0.5** NS 194.7 5/5 1 +toxic *ground Purina chow **Zn3P2 at 1.6% in EPA standard food and challenge bait consumption signi- ficantly different, P<0.05; NS=not significant TABLE XXVI Choice tests using lab rats ejqoosed to EPA baits containing 1% ejqoerimental zinc phosphide formulations and EPA placebo baits. Buffering Wash Mean Animal Body 3-Day Consumption t-test Mean Dose Mortality Mean Time to Substances Weight (g) (g) (X ± S.E.) (mg/kg) # Deaths/ to Death Initial Final EPA Toxic bait Sub. Lethal Test Animal (days) NaHCO3 + 260 264 30.±1.5 1.9±0.6 NS 30.6 78.5 4/5 1 NaHCO3 - 268 - 8.6+5.9 1.4+0.4 NS 17.6 57.2 4/5 1 . - + 255 255 14.3+12.3 1.8±0:4 NS 36.7 78.3 4/5 1 - - 265 267 8.6±5.2 2.2±0.5 NS 68.4 89.3 4/5 1 NS = toxic and challenge bait consumption not significantly different, P<0.05 00 65 rat was too small to degrade the acid condition in the stomach. C. Physical Characteristics, of Zinc Phosphide Commerical zinc phosphide may be less efficacious as a rodenti cide because it is rapidly decomposed by chemical reaction in the stomach of rodents after ingestion and produces toxic symptoms that may stop the animals from eating enough poisoned bait to receive a lethal dose. Zinc phosphide also has contact with sensory surfaces in the mouth of the rodent, which may increase the development of bait shyness through the gustatory sense. Many modifications of the physical formulation of zinc phosphide are possible, but the specific constructions made for the following evaluations cannot he detailed at this time. A variety of coded zinc phosphide formulations were prepared by Hooker Chemical company. These were evaluated in choice and non-choice tests with laboratory and wild rats and wild house mice. All formulations were tested at 1% in EPA-standard food. The results are summarized in Tables XXVII, XXVIII, and XXIII. Although discrimination by lab rats among the formulations occurred, complete mortality occurred in most cases, even when a choice of placebo bait was provided. Selected formulations were tested with wild rats and mice in a choice test regimen. Zinc phos phide formulation #1-21 gave but 20% mortality to wild rats, and only formulation # 7-14 provided total kill and the desired 100% TABLE XXVII Non-choice tests using rats (Rattus norvegicus) and mice (Mus musculus) exposed to EPA-standard food containing 1% experimental zinc phosphide formulations, n=5. Species Zn3P2 # Body Weight (g) Pre-test Consumption One-Day Consumption Mean Mortality Death Formulation Initial Final GPC* (g) (X ± S.E.) Toxic bait (g) (X±SE) mg/kg # Deaths/ (days) Test Animal Lab rats 7-14 216 215 20.8 + 2.7 3.3 ± 0.1 159.5 5/5 1 Lab rats 4-7 275 262 18.3 ± 3.0 1.8 ± 0.5 65.4 5/5 1 Lab rats 2-14 206 216 19.8 + 2.1 5.4 ± 0.7 262.4 5/5 1 Lab rats 7-11 213 215 19.5 + 1.4 4.0 ± 0.7 188.0 5/5 1 Wild rats 2-14 322 307 20.4 + 2.8 2.8 ± 0.5 97.9 5/5 1 Wild rats 7-11 298 281 21.0 ± 1.9 2.8 + 0.8 94.7 5/5 1 Wild mice 2-14 13 13 2.2 ± 0.4 1.2 ± 0.6 799.5 5/5 1 oo *Ground Purina Chow OV TABLE XXVIII Choice tests on lab rats with different Zn3P2 formulations at 1% in EPA and EPA placebo baits, N—5. Zn3P2 # Avg. Animal Body 3-Day Consumption t-test Mean Dose (mg/kg) Mortality Mean Time to Formulation Weight (g) (g) (X ± S.E.) # Deaths/ Death Initial Final EPA Toxic bait Sublethal Lethal Test Animal (day) 2-29 166 162 3.5±0.7 1.9+0.2 NS — 115.4 5/5 1 1-21 166 171 2.0±0.7 3.6+0.7 NS - 212.3 5/5 1 1-14 161 160 5.411.9 2.210.2 NS - 137.1 5/5 1 2-14 250 262 1.6+0.8 3.1+0.6 NS - 125.5 5/5 1 7-11 227 225 4.2±2.6 2.210.3 NS - 98.2 5/5 1 7-14 273 272 4.2+1.6 1.9+0.3 NS 74.6 5/5 1 6-10 163 174 1.5+0.5 2.410.4 NS - 147.3 5/5 1 4-7 258 251 6.2+3.8 0.8+0.1 NS 12.23 40.3 4/5 ' 1 4-6 166 173 4.011.5 1.7+0.3 NS - 100.1 5/5 1 4-0 153 178 26.0+12.9 1.210.4 + 65.18 92.5 3/5 1 NS = not significantly different + = significantly different (P£0.05) oo 88 mortality to wild rats. Though formulation 2-14 gave only 80% mortality to wild rats, it had the trend of highest toxic bait consumption (compared to other formulations). Zinc phosphide formulation #2-14 was selected for testing on wild mice. It provided only 20% mortality to this species, probably because of a nibbling behavior of wild house mice, as indicated by Jackson (1965), that enabled them to develop bait shyness by associating their illness with the last food they had eaten. D. Improvement of Zinc Phosphide Microencapsulations Microencapsulation has been used to improve the efficacy of roderiti- cides such as norbormide (Fall, 1978), alphachloralose (Greaves et al., 1968), zinc phosphide, and warfarin (Cornwell, 1971). Such a process attempts to mask the taste and odor of the chemicals; therefore baits will be more palatable to rodents and are less likely to elicit warning symptoms before the animals consume a lethal dose. Greaves et al. (1968) reported that gelatine-encapsulated norbormide (4:1 ratio of active compound to capsular material) in medium oatmeal increased the bait consumption and mortality significantly for lab rats. They also found that consumption of food containing ethylcellulose- encapsulated alphachloralose (10:1 ratio of active compound to capsular material) was increased significantly by mice, and mortality was consis tently greater with increasing concentrations of the poison. However, Cornwell (1971) reported that while encapsulated warfarin (6:1 ethyl- cellulose) at 0.5% concentration and encapsulated zinc phosphide (4:1 in ethylcellulose) at 1%, 2%, and 4% markedly improved the intake of the 89 toxicant, there was no significant increase in mortality. He suggested that the capsule wall was not sufficiently digested in the animal's gut to release the toxicant. Abrams and Hinkes (1974) also found that encap sulated warfarin was more acceptable than unencapsulated forms in identical baits and gave higher percentage of mortality to the rats. Ericsson et al. (1971) also reported that microencapsulated U-5897 significantly improved bait acceptance by rats (Rattus norvegicus) . The objective of this study was to evaluate the efficacy of various encapsulation formulations of zinc phosphide. The results are summarized in Tables XXIX and XXII* Seven out of 18 formulations provided 100% mortality to lab rats, and three of them (formulations 30-B, 25-B, and 30-D) also were correlated with a high degree of toxic bait consumption, more than 40% of the total food (EPA and toxic food) eaten. Of these, formulation 30-D was the most effective against wild rats, since it pro vided 80% mortality to the test animals, while the other formulations gave only 40% and 60% mortality. Furthermore,- it provided 60% mortality to the test animals when It was mixed with EPA standard food at 0.5%, while Zn3P2 combined with NaHCO3 (washed formulation) mixed with EPA standard food at the same concentration gave only 40% mortality to wild rats. In most cases encapsulated Zn3P2 baits killed lab rats within one day (except formulations 18-B and 25-G); in contrast, deaths of wild rats extended to two or three days (except formulation 18-D). Wild rats normally are slightly less susceptible to zinc phosphide than lab rats, and encapsulation apparently accentuates this difference. A portion of the toxicant may not have been released from the encapsula ting material, thus reducing the toxic impact; but lack of a repellent taste factor allowed the wild rats to continue feeding and to obtain a TABLE XXIX Summary of choice tests using lab rats exposed to EPA-standard food containing 1% experimental microencapsulated zinc phosphide, n=5. Microencapsulated Body Weight (g) 3-Day Consumption t-test Mean Dose Mortality Percent Mean Time Zn3?2 Formulation Initial Final (g) (X ± S.E.) (mg/kg) # Deaths/ Kill to Death EPA Toxic Bait Sub. Lethal Test animal (days) 30-A 406 384 15.216.9 2.310.6 NS 34.0 73.2 3/5 60 1 30-B 410 387 2.910.9 5.411.5 NS 57.6 145.4 4/5 80 1 25-A 401 372 11.011.5 3.711.1 + - 86.4 5/5 100 1 25-B 249 252 6.311.2 4.811.1 NS - 193.4 5/5 100 1 18-A 233 222 8.810.7 2.010.3 + - .. 83.3 5/5 100 1 18-B 249 238 8.712.8 2.711.5 NS - . 117.9 5/5 100 2 30-C 159 158 5.212.4 1.610.3 NS - 100.7 5/5 100 1 30-D 165 162 4.411.6 3.511.0 NS - 210.7 5/5 100 1 25-C 167 187 25.9110.2 1.710.2 + 74.4 119.8 3/5 60 1 . 25-D 163 182 12.618.2 2.210.5 NS 91.7 116.0 3/5 60 1 18-C • 169 196 26.419.6 1.910.2 + . 91.5 138.2 2/5 40 1 18-D 165 169 5.311.1 2.010.1 + - 123.7 5/5 100 1 30-E 160 200 18.7110.7 1.110.1 NS 70.9 57.7 1/5 20 1 25-F 158 180 5.612.7 1.410.2 NS 78.6 99.7 3/5 60 1 25-G 160 169 14.517.9 1.510.3 NS 51.0 107.0 4/5 80 2 18-F 158 185 17.616.4 1.310.5 + 51.9 128.3 2/5 40 1 18-G 154 176 13.815.2 1.410.2 + 92.9 79.9 3/5 60 1 10-F 159 175 24.5111.1 0.910.1 NS 59.2 56.8 3/5 60 1 «o o +- significantly different (P*Q.Q5). NS •’ not significantly different 91 lethal dose. This encapsulation allowed greater toxic bait consumption and increased efficacy of these formulations. E. Tests on Colonies of Wild Rats The objective of this study was to evaluate the efficacy of zinc phosphide baits under conditions allowing free access to baits and behavioral interactions. Four different encapsulated formulations of zinc phosphide were selected for choice tests on different colonies (n=5) of wild rats kept in metal tanks. The results are summarized in Table XXX. Two formulations (30-D and 18-D) provided 100% mortality to the test animals; the other two formulations (30-B and 2-14), only 40% mortality. This indicated that some microencapsulated Zn3P2 formula tions were more effective in killing wild rats than non-microencapsulated ones. In conclusion, microencapsulation increased the efficacy of Zn3P2 when tested on wild rats that were kept in confined populations (compared to the tests on those rats that were kept in individual cages). Micro- encapsulated Zn3P2 30-D was selected for further investigation in a field situation. III. Field Test The purpose of this study was to test the efficacy of microencapsula ted Zn3P2 30-D under field conditions. The test program was conducted outside a hog barn, as indicated in Figure 3. The weedy vegetation grew considerably during the program, becoming quite dense by the time of the post-treatment census. The predominant TABLE XXX Choice tests on colonies of wild rats (n=5) with experimental zinc phosphide at 1% in EPA and EPA placebo baits. Zinc phosphide Avg. Animal Body Total Daily Pre -test Total 3-Day Consumption Mortality Mean Time Formulation # Weight (g) Cbnsumption on Ground During Treatment (g) # Deaths/ to Death Initial Final Purina Chow (g) EPA Zn3P2 Bait Test Animal (days) Day 1 Day 2 Day 3 30-D* 332 282 35.8 49.5 58.7 26.0 12.0 5/5 2 18-D* 279 282 127.9 180.4 180.3 82.4 27.6 5/5 2 30-B* 256 264 111.4 147.7 154.8 222.6 14.9 2/5 3 2-14 242 229 98.1 123.0 134.8 230.9 6.6 2/5 2 V£> to ♦encapsulated 93 species were foxtail (Setaria sp.), thistle (Circium spp.), and crab grass (Digitaria sp.). In the dense cover there were several species of weeds, such as ragweed (Ambrosia spp.), milkweed (Asclepias spp.), button weed (Abutilon theophrasti), mustard (Brassica spp.), and smartweed (Polygonum spp.). Temperatures ranged from 10° C to 41° C. The average daily minimum and maximum were 20° C and 35° C, respectively. There was slight precipi tation on the second day of treatment period and heavy precipitation on the third day of post-treatment census. Bait consumption during pre-treatment census, treatment period, and post-treatment census is summarized in Table XXXI and shown in Fig. 6. Rodent activity (determined from rodent track indices) is shown in Fig. 7. During the pre-treatment census none of the food was taken from bait stations #3 and 5; and only small amounts were taken at bait stations #1, 2, and 11. The maximum food consumption on the 4th day (189 g) was due to the heavy consumption at bait station #12, which had been placed under dense weed cover on the third day. Rat activity on the first day of poisoning was almost equal to that of the last day of pre-treatment census; however, the amount of bait consumed by rodents on the first day of poisoning was much less than was consumed on the last day of the pre-treatment census. There was a decreas ing trend in both toxic bait consumption and rodent tracks during the initial six days of treatment. Some inverse rat/mouse activity was evident. On the third day when rat activity had decreased, the mouse track index doubled; and this same phenomenon occurred later in the treatment and post-treatment periods as TABLE XXXI Summary of bait consumption in field test. Bait Pre-treatment (4 days) Treatment (10 days)** Zn3P2 30-D at Post-treatment EPA Station EPA standard food 1% in hog food eaten by rodents (g) standard food eatei Number eaten by rodents (g) by rodents (g) 1 2 3 4 1 234567 8 9 10 1 2 3 4 1 0.0 1.0 0.0 0.0 9.0 17.7 5.0 0.0 0.0 0.0 0.0 0.0 0.5 0.0 2.1 14.5 20.6 67.8 2 0.0 0.0 2.8 1.0 0.0 0.0 0.0 0.0 0.0 1.5 0.0 34.8 0.0 0.0 0.0 0.0 0.0 0.0 3 0.0 0.0 0.0 0.0 0.5 0.0 0.0 1.5 2.0 0.0 11.5 3.5 0.0 26.3 0.0 0.8 2.2 66.7 4 4.5 2.0 6.0 2.0 0.5 5.0 2.8 0.0 1.6 0.5 0.0 0.0 0.0 0.3 3.1 2.6 5.9 124.0 5 0.0 0.0 0.0 0.0 0.5 0.0 2.2 2.3 1.2 1.5 0.2 0.5 0.5 0.4 1.5 1.0 1.1 24.9 6 2.0 4.0 37.0 34.7 0.5 1.0 0.0 1.5 3.5 0.0 1.1 0.0 0.5 1.8 28.0 7.3 10.5 60.3 7 7.0 10.5 9.5 9.4 0.0 0.0 1.0 3.5 0.7 0.0 0.0 4.8 32.2 46.5 0.0 1.0 22.4 46.5 8 33.0 32.0 21.5 12.3 3.2 0.0 1.8 2.5 1.0 1.2 1.8 0.0 1.6 1.9 2.1 4.6 5.1 12.5 9 12.4 17.0 8.2 11.0 2.8 0.5 1.5 3.0 1.0 0.8 0.0 0.0 2.3 3.3 6.5 5.6 7.8 17.3 10 4.5 8.5 7.0 4.0 0.5 2.0 0.0 2.0 0.0 0.0 4.9 0.0 1.0 3.3 14.0 14.7 12.2 15.3 11* ■ -. - 0.0 3.0 1.1 0.5 0.8 0.5 1.0 1.8 0.0 27.8 6.0 0.0 17.0 16.2 22.8 36.3 12* - - 75.0 111.6 7.8 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 1.8 10.3 13.9 99.6 Total 63.4 75.0 167.0 189.0 26.2 26.7 15.6 16.8 12.0 7.3 19.5 71.4 44.6 83.8 76.1 78.6 124.5 471.2 *Bait station was placed on the 3rd day of pre-treatment census **Total 10-day toxic bait consumption = 323.9 g l£> ■U 95 Figure 6. Bait consumption by outside populations of rats and mice at a hog farm near Waterville during 16 July through 8 August, 1978. Pre-treatment Treatment Post-treatment Census Census ) g ( N O I d T o P i M r U e S p N O C lag T I y A a B d - 3 97 Figure 7. Rat and mouse tracks on tracking boards (expressed in terms of index numbers) found during the efficacy field trial of Zn3P2 30-D. TRACK INDEX D A Y S t o 99 well. Characteristically mouse infestations are reduced in areas of heavy rat infestation, and vice versa, since mice do not compete readily with rats for food. Jackson (1972) similarly indicated that mouse pop ulations appeared to increase through the sanitation and poisoning programs in urban areas as rat populations decreased. By the end of the first week rodent activity had been reduced about 50%. Some of the surviving animals may have rejected the bait initially (in favor of untreated food abundantly available in the environment) or developed bait shyness following consumption of sublethal doses. After the first week of toxic bait exposure, both feeding and track activity increased, suggesting the invasion of peripheral animals into the biological vacuum created by the mortality of resident animals (Calhoun and Webb, 1953; Calhoun, 1962). The feeding and tracking acti vity at bait station #7, which was placed in the middle of dense grass cover (area B in Figure 3) during the treatment period, pointed to such movements. The toxic bait consumed by the rodents at this bait station was reduced to 0.0 g during the sixth and seventh days of the poisoning program but it increased from 4.8 g on the eighth day to 46.5 g on the last day of the treatment period. Though rat and mouse trails near this bait station were not evident, rodents could gain access from all directions. The toxic bait consumption at bait station #11, which was placed near the dense weed cover (area C in Fig. 3), also indicated movements. For instance, the toxic bait consumption at this station had dropped to 0.0 g on the seventh day of the poisoning program but increased to 27.8 g on the following day. In several cases bait con sumption dropped sharply after a day of heavy consumption, suggesting 100 that these consumers had died, were sick but subsequently died, or were bait shy. This study allowed a 3-day lag period after the poisoning program to counter bait shyness or position effects, but this aspect may have been countered by movements of animals. If the rats and mice that lived very close to the bait stations had been killed by the poison, those rats and mice inhabiting peripheral habitats and not previously ejqoosed to the toxic baits might have invaded the baiting area. The consistent increase of bait consumption at all bait stations during the post-treatment census reinforced this assumption. Alternatively, the increase in consumption could mean that the residents were learning locations of stations or overcoming their neophobia, although this hypothesis seems less appro priate because of the long period of bait exposure. The highest daily consumption in post-treatment census was 471 g, which was a i49% increase compared to the highest daily consumption of pre-treatment census. A census technique using tracking boards also revealed increased activity patterns. Rat tracks (expressed in index values) in the post-treatment census increased 12% (in comparison to the highest daily index in the pre-treatment census), and there was a corresponding 12.5% reduction of mouse tracks. In this study 324 g of toxic bait were taken by rodents. An estima ted 135 rats would be expected to have been killed (based on lab results that had shown 12 g killed five wild rats, Table XXX) „ However, only two rats and two mice were found above ground during the treatment period; many rats and mice probably died in their burrows. In the post-treatment census trapping program 13 rats (6 males and 7 females) and six mice (2 males and 4 females) were captured in three 101 nights. Of these, four were young rats (weighing less than 100 g); all the rest were adult rats and mice. Though total reduction of the rat and mouse populations were not obtained in this study, microencapsulated zinc phosphide 30-D was found ■ < ' to be accepted by these rodents. If the baiting program had been extended both in time and space until the total population had been exposed to the toxic bait, a greater measure of control likely would have been achie ved. In other studies, Twigg (1962) used zinc phosphide at 2.5% and 5.0% on cereal baits (e.g., paddy, broken rice without husk, and a mixture of broken rice and maize) together with arsenious oxide and warfarin baits for controlling marsh rat (Rolochilus sciureus) in sugarcane fields in British Guiana. He found that about 50% of the rodent population still remained after the baitings. Kapoor and Khare (1965), using zinc phos phide at 5% level on Pakoras (made out of gram and vegetable) for control ling the black rat (Rattus rattus) in the rice mill at Saharunpur, reported 96% reduction of the rat population. However, West et al. (1972) failed to reduce damage by the ricefield rat (R. r. mindanensis) to the paddy field by use of zinc phosphide at 1% on polished rice combined with coco nut oil at 0.5% by weight. They indicated that poor initial acceptance might be a factor that reduced efficacy. Hilton et al. (1972b)tested the efficacy of zinc phosphide on Polynesian rats (R. exulans) under labora tory conditions and found that a 2% zinc phosphide bait on oat groats provided 100% mortality to this species with a high acceptance ratio (compared to other concentrations of Zn3P2 on the same bait material). One of their studies under field conditions revealed that after applying 102 2% Zn3P2 on oat groats at 5 lb/acre by aerial application in unprebaited fields of 16-month-old sugarcane, the rat population was reduced about 70%; about 87%, in the fields that were prebaited with untreated oats 10 days before toxic bait application. Dubock and Rennison (1977) found no significant differences in level of control achieved among several cereal baits tested. 103 SUMMARY The efficacy of zinc phosphide when applied for rodent control often has been considered reduced due to poor initial bait acceptance and devel opment of bait shyness. This study investigated^the bait shyness pheno menon, modified forms of zinc phosphide, acceptance of various baits under laboratory conditions, and testing of a selected zinc phosphide formulation in the field. Wild Norway rats, but not lab rats, developed bait shyness to zinc phosphide bait after surviving a sublethal dose and avoided consuming toxic baits on subsequent exposure. The shyness was largely to the zinc phosphide rodenticide, but not to the specific bait material. Some 3 ' wild, bait-shy rats retained memory for zinc phosphide for at least 60 days. Social behavior of adult rats in a small colony had no influence on bait shyness development in the members of the group. In food preference studies, red pepper at 0.16%, coconut flakes at 1.25%, 2.5%, and 5%, and brown or granulated sugar at 10% levels signifi cantly increased palatability of EPA-standard food for lab rats. Of these only red pepper at 0.16% increased palatability of EPA-standard food for wild rats, but it was rejected by wild rats when mixed with a toxic bait containing Zn3 P2. Bait combinations A (consisting of ground barley and rolled oats), B (consisting of ground wheat and rolled oats), and E (consisting of ground barley, ground corn, and rolled oats) were preferred to EPA-standard food by lab rats; but wild rats showed no preference for these combinations over EPA-standard food. The addition of buffering substances to Zn3P2 did not significantly improve the toxic bait consumption. All the special formulations provided 104 100% mortality to lab rats in non-choice feeding situations. In choice regimens, complete mortality occurred to lab rats in most cases, but only formulation #7-14 resulted in 100% mortality to wild rats. Though formu lation #2-14 provided only 80% mortality to wild rats, consumption of this toxic bait was higher than with the other formulations evaluated. Seven microencapsulated zinc phosphide formulations provided complete mortality to lab rats in choice tests. Of these, microencapsulated for mulation #30-D gave 80% mortality to wild rats, while the other microen capsulated formulations provided only 40% to 60% mortality. In colony tests with wild rats microencapsulated zinc phosphide formulations #30-D and #18-D provided 100% mortality, while a comparable unmicroencapsulated formulation #2-14 gave only 40% mortality. In a field trial microencapsulated zinc phosphide #30-D was well- accepted by rats and mice. One hundred and thirty-five rats were estimated to have been killed from this poisoning program; however, the rodent population was not reduced significantly because of immigration from surrounding environments. The development of special zinc phosphide formulations and use of microencapsulation improved the efficacy of zinc phosphide baits. However, more research and evaluation is needed before the introduction of these compounds for efficacious rat control in field situations is possible. ANNOTATED BIBLIOGRAPHY ZINC PHOSPHIDE AS A RODENTICIDE by Kasem Tongtavee from Zinc Phosphide Development for Rat Control Ph.D. Dissertation Bowling Green State University 1978 < * 105 ANNOTATED BIBLIOGRAPHY Abrams, J. and T.M. Hinkes. 1974. Acceptability and performance of encapsulated warfarin. Pest Control 42(5):14-16. Reports the tests of lab and wild rats for the efficacy of encapsulated warfarin, indicates that an encapsulated warfarin is more acceptable than unencapsulated warfarin in identical baits. Encapsulated war farin also increases percentage of mortality to the tested animals. Andreev, S.B., A.V. Voevodin, V.A. Molchanova, and A.V. Khotyanovich. I ' ' 1958. Some results of the use of tracer techniques in the study of plant protection. (Transl. from Russian). Proc. 2nd U.N. Internat. Conf. Geneva 27:85-92. Reports the results of studies concerning use of radioisotopic method in tackling the problems of plant intoxication and insect intoxicol ogy. Pathway of zinc phosphide administered to lab rats also is determined by this procedure. Anon. 1963. Zinc phosphide. Hooker Chem. Co. Data Sheet No. 809. 2 p. Briefly describes the properties of zinc phosphide and suggests use of this compound as a rodenticide for rats and other rodents. Anon. 1967a. Tartar emetic, its use and evaluation. National Pest Control Association Tech. Rel. 4-67. 2 p. Suggests use of tartar emetic in zinc phosphide bait as a safety agent in the case of bait ingestion by cats and dogs. However, it usually decreases the acceptance of the bait by rats; so it is not used frequently today. . 106 Anon. 1967b. Zinc phosphide. National Pest Control Assoc. Tech. Pel. 17-67. 6 p. Physical and chemical properties, stability, toxicology, and hazards of zinc phosphide are described. Bait preparations and application of poisoned baits also are explained. Anon. 1968. Research report on the stability of zinc phosphide. National Pest Control Association Tech. Rel. 14-68. 2 p. The decomposition of zinc phosphide dusted onto wet baits (i.e., apples, potatoes) is reported. Care in the placement of zinc phos phide baits also was suggested. Anon. 1975. Rodenticides: analysis, specifications, formulations for use in public health and agriculture. Report of an Informal Consul tation of Experts to WHO and FAO, WHO VBC/76.3, 76 p. Physical and chemical properties of zinc phosphide être indicated. Zinc phosphide is used in baits at 0.75 to 5% concentration for controlling rodents. Anon. 1976. Zinc phosphide. National Pest Control Association ESPC: 041426-041427. Chemical properties and toxicity of zinc phosphide are discussed. Antidote for zinc phosphide poisoning is suggested. ASTM. 1961. Standard specifications for sieves for testing purposes. ASTM:Ell—61, 1479-1486. These specifications cover wire cloth sieves, round-hole plate screen (sieves), and square-hole plate screens (sieves) for precision 107 testing in the classification of materials according to size (mech anical analysis, fineness, and particle size determinations). ASTM. 1977a. Standard Test Method for Efficacy of Acute Mammalian Preda- cides. ASTM Standards on Vert. Control Agents E552-75;6-10. Standard test method for efficacy of acute mammalian predacides under laboratory conditions and initial field trial is suggested. ASTM. 197'^. Standard test method for efficacy of a single-dose rodenti cide under laboratory conditions. ASTM standards on Vert. Control Agents E565-76;16-23. Standard test method for efficacy of a single-dose rodenticide under laboratory conditions is suggested. Barnett, S.A. 1956. Behavior components in the feeding of wild and laboratory rats. Behaviour 9:24-43. Reports the study of feeding behavior of rats and concludes that 1) wild-type rats tended to prefer grains to meal, 2) both wild and albino rats showed very marked exploratory and sampling behavior which enabled rats to change rapidly from a familiar distasteful food to an unfamiliar but palatable one, and 3) wild rats, but not albinos, tended to develop flight and avoidance behavior. Harnett, S.A. 1959. Experiments on "Neophobia* in wild and laboratory rats. Brit. J. Psychol. 49(3):195-201. Reports that all rats (R. norvegicus), wild or tame, have a tendency to explore their surroundings. Wild rats, but not tame ones, also tend to avoid unfamiliar objects in their familiar surroundings. The placing of a new object near or at a rat's usual feeding place may lead to the reduction of the rat feeding. • 108 Barnett, S.A., P.E. Cowan, G.C. Radford, and I. Prakash. 1975. Peripheral anosmia and the discrimination of poisoned food by Rattus rattus L- Behavioral Biology 13:183-190. Reports the ability of Rattus rattus to discriminate zinc phosphide bait from non-toxic food after having survived a sublethal dose from stomach tube administration. The results of making rats anosmic by treatment of the nasal mucous membrane with a solution of zinc sul phate (ZnSO0 also are discussed. ' Bentley, E.W., I»JE. Hammond, A.H. Bathard, and J.H. Greaves. 1961. Sodium fluoroacetate and fluoroacetamide as direct poisons for the control of rats in sewers. J. Hyg. Camb. 59:413-417. Reports the field trials for rat control in sewers in Surrey, Great Britian, and suggests that three monthly operations against rats in sewers using either 0.25% sodium fluoroacetate or 2% fluoroacetamide are more effective than six monthly treatments with 2.5% zinc phosphide. Bhardwaj, D. and J.A. Khan. 1978. Effect of texture on the food prefer- enees of bait-shy wild rats (Rattus rattus L.). II. Proc. Indian Acad. Sci., 87B (Animal Science) (3):77-80. Black rats (Rattus rattus L.), poisoned with 0.02% zinc phosphide in lidióle lentil (Lens esculenta) and green gram (Phaseolus aureus), do not accept these whole grains again. However, they do not avoid the alternative forms of the same foods (e.g., husked lentil and husked green gram. Use of alternative forms of the same foods as 0 . . the bait carriers of Zn3P2 is suggested for poisoning this species. Blaxland, J.D. and R.F. Gordon. 1945. Zinc phosphide poisoning in 109 poultry. Vet. Jour. 101:108-110. Reports the study of toxicity of zinc phosphide to poultry; states that it is an extremely toxic compound for poultry. Brock, E.M. 1965. Toxicological feeding trials to evaluate the hazard of secondary poisoning to gopher snakes, Pituophis catenifer. Copeia 2:244-245. Evaluates the reaction of gopher snakes (Pituophis catenifer) which had ingested small rodents (i.e., Microtus californicus, Peromyscus maniculatus) that had consumed various amounts of toxicants, such as sodium fluoroacetate, strychnine alkaloid, endrin, arsenic tri oxide, zinc phosphide, thallium sulfate, and three anticoagulants (Prolin, warfarin, and diphacin). ' Brooks, J.E. 1962. Method of sewer rat control. Proc. 1st Vert. Pest Control Conf., California:227-244. Discusses general aspects of sewer rats and explains use of poisoned baits (i.e., sodium fluoroacetate, fluoroacetamide, Zn3P2, warfarin, Pival, and Fumarin) for sewer rat control. Use of paraffin bait as moisture-proofing also is suggested. Brooks, J.E. and A.M. Bowerman. 1973. Preferences of wild Norway rats for grains, seeds, and legumes. Pest Control 41:13, 16, 18, 36, 38-39. Discusses different kinds of grains that were successfully used as bait materials for rodenticides in rat campaigns. . The results of testing for preferences of wild rats in a choice between cornmeal and another food (e.g., grains, seeds, and legumes) are reported. Bull, J.O. 1972. The influence of attractants and repellents on the no feeding behavior of R. norvegicus. Proc. 5th Vert. Pest Conf., CA: 154-160. Describes a rational evaluation of attractants and the influence of different odors in inducing R. norvegicus to feed at given locations. The influence of certain repellents (i.e., undiluted extracts of Latakia tobacco) also is examined. Calhoun, J.B. 1962. Population density and social pathology. Sci. Amer. 206(32):139-146. Reports study of population density of rats in a confined space provi ded with abundance of food and dwellings. Social activities and ab normal behavior of overcrowded rat populations also are discussed. Calhoun, J.B. and W.L. Webb. 1953. Induced emigrations among small mam mals. Sci. 117(3040)-.358-360. Reports the study of the effect of intensive trapping over a large area upon the members of the surrounding population. The movement of small mammals (e.g., Peromyscus, Clethrionomys, Blarina and Sorex) into a study area after trapping are discussed. Chitty, D. 1954. Control of rats and mice. Oxford Univ. Press, Vol. 1:305 p. Comprehensive; explains various methods for controlling rats and mice. Hazards to man and domestic animals also are reported. Chitty, D. and M. Shorten. 1946. Techniques for the study of the Norway rat (Rattus norvegicus). J. Mammal. 27:63-78. Reports the observation of feeding behavior of the Norway rats, indi cates that there is great individuality about the way different rats approach their food; some rats settle down in one place and feed for Ill 1-50 minutes, while others seize a mouthful of food (e.g., wheat) and run away to eat it under cover. The avoidance behavior of rats to unfamiliar objects also is discussed. Clark, D.O. 1972. The extending of cotton rat range in California: their life history and control. Proc. 5th Vert. Pest Conf.:7-14. Reports damages done by cotton rats (Sigmodon hispidus) on varieties of field crops. Habitats and life history of cotton rats also are described. Zinc phosphide and strychnine baits are suggested for control program. Clark, D.O. 1975. Vertebrate pest control handbook. Calif. Dept. of Food and Agri.:522-1 - 522-7. . Discusses biology and legal status of ground squirrels (Spermophilus beechegi, S. beldingi beldingi, and S. beldingi oregonus). He also recommends use of oat groats coated with zinc phosphide at 1% for spot baiting and at 2% for broadcast baiting in controlling ground squirrels. Cooper, P. 1962. Poisoning by drugs and chemicals. Charles C. Thomas, Springfield, Ill., 264 p. Discusses the properties of zinc phosphide, the action of phosphine gas, and the symptoms of zinc phosphide poisoning. Cornwell, P.B. 1971. New agents of.control - microencapsulation of rodenticides. Proc. of 3rd British Pest Control Conf.:42-51. Reports the results of using microencapsulation for improving the efficiency of norbormide and zinc phosphide in killing albino mice 112 and albino rats and indicates that microencapsulation increases the acceptance of the bait by rats and mice but higher mortality cannot be obtained. Cornwell, P.B. and J.O. Bull. 1967. Taste preferences in rodenticide development. Pest Control 35(8):15, 16, 18, 20, 64. Reports the results of testing lab and wild rats (Rattus norvegicus) for food preferences in a choice situation, indicates that the rats showed preference of grains in the following order: maize, wheat, and rice. A 50/50 mixture of oat and maize meal is as acceptable as oatmeal alone. Crabtree, D.G. 1962. Review of current vertebrate pesticides. Proc. 1st Vert. Pest Control Conf., California:327-362. Discusses various aspects (i.e., physical properties, physiological action, and limitations) of vertebrate pesticides (i.e., strychnine, sodium monofluoroacetate, thallium sulfate, zinc phosphide, cyanide, red squill, ANTU, warfarin, and diphacinone). The use of tracking powder containing toxic substances, such as arsenic trioxide and en drin, for commensal rodent control also is ejqolained. i ■ Crabb, W.D. and L.D. Emik. 1946. Evaluating rat baits by field acceptance trials on Guam. J. Wildl. Mgmt. 10:162-171. Reports the study of food preference of the Polynesian rat Rattus exulans and the black rat Rattus rattus in field situations on Guam, indicates that rolled oats appear to be the cereal choice (as com pared to wheat). Three other bases are superior as adjuvants to the cereal in the following order: ground fresh copra, copra oil, and 113 peanut butter. The optimum ratio of copra to rolled oats is found to be 1:4 to 1:6. ' Curry, A.S., D.E. Price, and F.G. Tryhorn. 1959. Absorption of zinc phosphide particles. Nature 184(4688):642-643. Reports the results of using zinc phosphide labelled with ^2P to demonstrate particles of this compound in the livers of the poisoned rats. He indicates that zinc phosphide particles, not phosphine gas, cause damage to liver tissues. Dana, R.H. 1962. Ground squirrel control in California. Proc. 1st Vert. Pest Conf.:126-143. Discusses the use of chemicals, fumigants, and trapping methods for controlling ground squirrels. Major poison baits are 1080, strych nine, thallium sulfate, zinc phosphide, and anticoagulants. Zinc phosphide bait commonly is used at 1% on whole barley or oat groats. Deoras, P.J. 1972. Rat reduction with indigenous methods. Proc. 5th Vert. Pest Control Conf., California:168-177. Discusses damage done by rats in India and reports that zinc phos phide in the form of paraffin capsules killed many rats in the rat campaign. Dorrance, M.J. and B.K. Gilbert. 1977. Consideration in the application of aversion conditioning, p. 136-144 IN: Test Methods for Vertebrate Pest Control and Management Materials, Jackson, W.B. & R.E. Marsh, eds. ASTM STP 625 p. Comprehensively discusses learned taste aversion of animals (i.e., rats, coyotes, and bears) and reports the case of successful use of 114 a drug, lithium chloride, for prevention of damage to bee hives by bears. Doty, R.E. 1945. Rat control on Hawaiian sugar cane plantation. Hawaiian Planter’s Record 49:71-231. Various aspects in biology and ecology of rats on Hawaiian sugar cane fields are discussed. Various control methods (e.g., use of fumigants use of virus, habitat modification,;and use of poisons) are also explained. The comparative efficiency of rat poisons (e.g., thallium sulfate, barium carbonate, strychnine, red squill, yellow phosphorus, and zinc phosphide) also is reported. Dreisbach, R.H. 1963. Handbook of poisoning, 4th edition. Lange Medical Publications. Los Altos, California: 467 p. Toxicity of phosphine gas is discussed. Treatment for- phosphine poisoning is suggested. Dubock, A.C. and B.D. Rennison. 1977. Field evaluations of baits for poison treatments against Norway rats. Proc. Brit. Crop Prot. Conf. -Pests and Diseases:451-460. Stabilized medium oatmeal and six other relatively simple cereal baits for rats were compared, both unpoisoned and poisoned, with 2.5% zinc phosphide in a field trial. Though greater weights of soaked wheat were eaten than of any other baits, there was no . significant difference in the level of control achieved using any of the baits Ce.g., barley meal, maize meal + barley meal (1:1), wheat + corn oil, stabilized medium oatmeal, wheat meal, maize meal, + thirds ;(1:2), and soaked wheat]. 115 Durairaj, G. and B.K. Guruprasad. 1975. Evaluation of rodenticides in the paddy fields at Negenahalli (Mysore district). All India Rodent Seminar, Ahmedabad, India:258-264. Discusses the use of rodenticides (i.e., aluminum phosphide, zinc phosphide, Fumarin, and warfarin) for controlling the Indian mole rat (Bandicota bengalensis) in paddy fields. The relative efficacy of each rodenticide also is reported. Zinc phosphide was used at 2% in a bait (consisting of 80% rice flour, 10% roasted groudnut, 10% ground nut oil) and reduced rat activity 69.2%. EPA. 1977. Standard test method for field evaluations of the efficacy of lethal rodent baits. Unpublished paper, 10 p. This method is based upon the use of pre- and post-treatment rodent censuses to assess the degree or percentage of rodent control achieved as the result of a rodent bait application. Methods of experimental design, site selection, census, treatment, and data analysis are sug gested. Parameters of efficacy also are indicated. Elmore, J.W. and F.J. Roth. 1943. Analysis and stability of zinc phos phide. J. Assoc. Office. Agri. Chem. 26:556-564. Reports the analysis of the stability of zinc phosphide when used in the form of poisoned grains under conditions of both storage and field use. The decomposition of zinc phosphide when exposed to acid and air also is indicated. Ericsson, R.J., H.E. Downing, R.E. Marsh, and W.E. Howard. 1971. Bait acceptance by rats of microencapsulated male sterilant alpha- chlorhydrin. J. Wildl. Manage. 35(3):573-576. 116 U-5897 induces a permanent lesion in the caput epididymidis of domestic and wild rats (Rattus norvegicus) with subsequent sterility. Bait with 1% concentration is sufficient to cause sterility or death, though food intake will be reduced. Microencapsulation of U-5897 significantly (P<0.01) improves bait acceptance. Evans, J. 1970. Methods of controlling jackrabbits. Proc. 4th Vert. Pest Control conf., California:110-115. Discusses damage to agricultural crops and rangeland done by the jackrabbit (Lepus californicus) and suggests use of 0.75% zinc phosphide carrot bait for controlling them. Fall, M.W. 1978. The behavior of confined populations of rats (Rattus norvegicus) with observations on their responses to baiting routines using the acute rodenticide, Norbormide. Ph.D. Thesis, Pennsylvania State University, Pennsylvania, 142 p. . A comprehensive study of the behavior of wild rats in confined pop ulations in responding to baiting routines using the acute rodenticide, Norbormide. The researcher indicates that agonistic interactions of the rats, particularly in the vicinity of the bait sources, pro bably is one of the major factors that reduces the efficacy of the acute rodenticide, and suggests reducing the agonistic interactions of the rats to improve the results of rodent control. Fitzwater, W.D. and I. Prakash. 1973. Handbook of Vertebrate Pest Control. Indian Council of Agri. Research, New Delhi, 92 p. Suggests the use of zinc phosphide at 2% in cracked corn or steam- crushed oats for controlling field mice; at 1.5% in pearl-millet, 117 sorghum, or Bengal-gram for controlling gerbils. Toxicity and antidote for this compound also are ejq>lained. Gaffrey, Q. 1953. The problem of odor attraction of rats. (In German) Anz. Scheldlingsk. 168-169. (Biol. Abstr. 30:29188). Observations oh Rattus norvegicus showed that odoriferous attractants appear to give more negative than positive results in rat control. However, zinc phosphide was not rejected by R. rattus when repeated after a sublethal dose. Gaines, T.B. and W.J. Haynes. 1952. Bait shyness to ANTU in wild Norway rats. Publ. Hlth. Repts. 67(3):306-311. Reports the results of testing wild Norway rats with ANTU for deter mination of bait shyness; indicates that wild Norway rats are bait shy at subsequent exposure to poison experienced earlier. Galef, B.G., Jr. and M.M. Clark. 1971. Social factors in the poison avoidance and feeding behavior of wild and domesticated rat pups. J. Comp. Physiol. Psychol. 75:341-357. . Reports the results of studies for poison avoidance and feeding behavior of wild and domesticated rat pups, indicates that rat pups born to colony members learn cues associated with food by following their parent to food, do not eat the diet that adults avoid, and continue to avoid that diet even after being isolated from their adults in a new environment. Garcia, J. and R.A. Koelling. 1966. Relation of cue to consequence in avoidance learning. Psychon. Sci. 4:123-124. Reports the results of testing lab rats with reinforcers (e.g., 118 bright-noisy water, radiation, lithium chloride, and delayed shock) in inducing learned taste aversions; indicates that all reinforcers are effective in producing aversions in lab rats. Garcia, J., F.R. Ervin, and R.A. Koelling. 1966. Learning with prolonged delay of reinforcement. Psychon. Sci. 5:121-122. Reports the study of gustatory aversions induced in lab rats. Apomor phine, a drug which causes nausea and emesis in humans, was found to induce taste aversion in lab rats. Garcia, J., F.R. Ervin, and R.A. Koelling. 1967. Bait-shyness: a test for toxicity with N=2. Psychon. Sci. 7:245-246. Reports the study of bait shyness developed by lab rats. It was found that cyclophosphamide (which produces diarrhea) induced taste aversion in lab rats. Garcia, J., W.G. Hankins, J.H. Robinson, and J.L. Vogt. 1971. Bait shyness: tests of CS-US Mediation. Physiology and Behavior 8:807-810 Reports the test of lab rats for bait shyness and the prolonged CS- US interval; indicates that when eating either wet or dry food is followed by illness rats will consume smaller amounts of the food on subsequent presentations. They also indicate that the gustatory system cannot account for the long-delay learning. In some of their experiments sweet water (1.0 g/L saccharin) or salty water (7.9 g/L sodium chloride) are used as taste cues, since both fluids are preferred to tap water by rats. Gratz, N.G. 1973. A critical review of currently used single-dose rodenticides. Bull. Wld. Hlth. Org. 48:469-477. 119 Briefly discusses the main characteristics of the single-dose rodenticides (i.e., ANTU, arsenic, trioxide, barium carbonate, norbormide, phosphorus, red squill, 1080, fluoroacetamide, stry chnine, thallium sulfate, zinc phosphide, crimidine, RS-150, L-chloralose). Recommendations for use of these rodenticides also are explained. Greaves, J.H. 1966. Some laboratory observations on the toxicity and acceptability of norbormide to wild rats (Rattus norvegicus) and on feeding behavior associated with sublethal dosing. J. Hyg. Camb. 64:275-285. This paper describes an acute toxicity assay of norbormide and zinc phosphide. The responses of rats that survived a sublethal dose.to a subsequent poison also are explained. Greaves, J.H., F.P. Rowe, R. Redfern, and P. Ayres. 1968. Microencap sulation of rodenticides. Nature 219(5152):402-403. Reports the use of microencapsulation to improve the efficiency of rodenticides by taste-masking or delaying the onset of their toxic effects. Norbormide and alphachloralose are the compounds used for this study. Greaves, J.H., M.A. Choudry, and A.A. Khan. 1977. Pilot rodent control studies in rice fields in Sind, using five rodenticides. Agro Ecosystems 3:119-130. Reports pilot rodent controls in rice fields in Sind, Pakistan, using five rodenticides (0.0375% coumatetralyl, 2.5% zinc phosphide, 0.03% sodium fluoroacetate, sodium cyanide powder, and aluminum phosphide tablets. Rodent species that caused damage to ¿eld crops 120 also are reported. Guerrant, G.O. and J.W. Miles. 1969. Determination of zinc phosphide and its stability in rodent baits. J. Ass. Off. Agri. Chem. 52(3): 662-666. Reports the study of zinc phosphide and its stability in rodent baits (i.e., 1% Zn3P2 apple and potato baits) by refluxing Zn3P2 with hydrochloric acid with subsequent determination by the alkali- metric quimociae phosphorus method. Hamar, M. 1976. Mechanized control of murids in isolated loci. Joint FAO/WHO/EPPO Conference on Rodents of Agri, and Public Hlth. Concern. Geneva, Switzerland:54-55. Discusses a mechanized method for the control of Mus musculus spici- legus, Apodemus sglvaticus, Apodemus agrarius, and Rattus norvegicus using zinc phosphide bait at 3% for protection of corn and sunflower crops on the Great Isle of Braila. Hankins, W.G.T J. Garcia, and K.W. Rusiniak. 1973. Dissociation of odor and taste in bait shyness. Behavioral Biology 8:407-419. Reports the tests of odor and taste in food-illness aversions of laboratory rats. The role of olfaction in the neophobic reaction to new foods also is discussed. Hargrave, G.E. and R.C. Bolles. 1971. Rat’s aversion to flavors following induced illness. Psychon. Sci. 23:91-92. This study reports the results of the tests on lab rats for the association of taste with illness. The rats were made ill by an intraperitoneal injection of 15% saline solution (3 ml/kg body weight). 121 Hausman, M.F. 1933. The behavior of albino rats in choosing food. II. Differentiation between sugar and saccharin. J. Comp. Psychol. (15):419-428. Reports the results of study of the behavior of lab rats in choosing food, indicates that rats prefer sugar solutions and saccharin solu tions to pure water. Hayes, J.W. 1975. Zinc phosphide. IN Toxicology of Pesticides. Bal timore: Williams and Wilkins. 580 p. Explains properties and toxicity of zinc phosphide. Effects of this rodenticide on domestic animals and wildlife also are described. Hayne, D.W. 1951. Zinc phosphide: its toxicity to pheasants and effect of weathering upon its toxicity to mice. Mich. Agri. Exp. Sta. Quart. Bull. 33(4):412-425. Zinc phosphide is reported to be highly toxic to pheasants with an average lethal dose of 9 mg/kg. Cracked corn bait carrying 2% zinc phosphide showed deterioration of toxicant after weathering for 27 days. Helal, T.Y., A.M. Ali, M.S. Arafa, A.M. Salit, and A.M. Abd-El-Wahab. 1975. Susceptability of Egyptian rodents to zinc phosphide. All Rodent Seminar, Ahmedabad, India. 248-252. Reports the tests of zinc phosphide-on several species of Egyptian rodents (i.e., Rattus norvegicus, R. r. alexandrinus Geoffory, R. r. frugivorus Rafinesque, Arvicanthus niloticus Desmarest, and Acomys cahirinus E. Geoffory and St. Hilaire) and indicates LD50 for all groups. 122 Hilker, D.M., J. Hee, H. Higashi, S. Ikehara, and E. Paulsen. 1967. Free choice consumption of spice diets by rats. J. Nutri. 91:129 131 • Reports the results of testing lab rats for food preference with spiced diets containing various species (e.g., ginger, cloves, cinnamon, black pepper) at concentrations ranging from 0.05 to 5% and non-spiced diets; indicates that adult rats do not show prefer ence between spiced diet and unspiced diet, while young rats prefer unspiced to spiced diets in choice situations. Hilton, H.W. and W.R. Robison. 1972. Fate of zinc phosphide and phos phine in the soil-water environment. J. Agr. Food Chem. 20(6): 1209-1213. Discusses the analysis of the stability of Zn3P2 in soil.and water. The hydrolysis of Zn3P2 at room temperature also is reported. Hilton, H.W., W.H. Robison, A.H. Teshima, and R.D. Nass. 1972. Zinc phosphide as a rodenticide for rats in Hawaiian sugarcane. Proc. of 14th Congress, ISSCT:561-570. Reports the investigations of toxicity and residue of zinc phosphide in the laboratory and in the Hawaiian sugarcane field. Rat habits, bait acceptance, and efficacy also are described. Hood, G.A. 1972. Zinc phosphide - a new look at an old rodenticide for field rodents. Proc. 5th Vert. Pest Control Conf., California:85-92. Describes toxicity, mode of action, and hazards of zinc phosphide and suggests the use of zinc phosphide as a rodenticide. 123 Howard, W.E. 1959. How to overcome bait shyness in rodents. Pest Con trol 27(8):9-13. Reports a case of bait shyness developed by lab rats to 1080 after having survived a sublethal dose and suggests using durable baits and prebaiting to overcome bait shyness in rodents. Howard, W.E., R.E. Marsh, and R.E. Cole. 1977. Duration of associative memory to toxic bait in deer mice. J. Wildl. Manage. 41(3):484-486. Reports that deer mice (Peromyscus maniculatus) that had been condi tioned by sublethal dose to avoid eating oat kernels coated with compound 1080 could retain their aversion to compound 1080 bait more than eight months; suggests that deer mouse control should not employ the same position at intervals shorter than eight months. Ingram, P.L. 1945. Zinc phosphide poisoning in a colt. Vet. Rec. 57 (9):103-104. Reports the primary hazard of zinc phosphide rodent bait to a 14 month-old colt in England and suggests the application of such a highly toxic product with the utmost care. Jackson, W.B. 1965. Feeding patterns in domestic rodents. Pest Control 33(8):12-13, 50, 52. Discusses feeding behavior of rats and mice; emphasizes the behavior patterns (e.g., exploration, avoidance, and flight) of wild rats and sampling behavior of house mice. The relationship of feeding pat terns to population density also is discussed. Jackson, W.B. 1972. Biological and behavioral studies of rodents as a basis for control. Bull. Wld. Hlth. Org. 47:281-286. 124 Discusses the growth dynamics of domestic rodent populations; ex plains the use of various control operations (i.e., trapping, poi soning, predation, and chemosterHants) against rats and mice. Determination of the movement patterns, the home ranges, and the behavior patterns of rodents also is suggested to achieve control in programs. Jackson, W.B. 1974. Use of microencapsulation to enhance rodenticide acceptance. Controlled Released Pesticide Symposium. Univ. of Akron:6.1-6.10. Comprehensively discusses the "ideal" rodenticide; suggests the use of microencapsulation to enhance rodenticide acceptance. Jackson, W.B. 1976. Phosphorus poisoning. J. Am. Med. Assoc. 236(8): 918-919. Discusses the proper use of zinc phosphide bait for rodent control and differentiates it from yellow phosphorus. Addition of emetic into rodent bait, as a safety factor in case of accidental ingestion by children or pets is indicated. Johnson, H.D. and E. Voss. 1952. Toxicological studies of zinc phosphide. J. Amer. Pharm. Assoc. 41(9):468-472. Reports the investigations of chronic effects of zinc phosphide on lab rats having consumed a sublethal dose of this compound for certain periods. Investigation of acute effects on other animals (e.g., dogs, cats, rabbits, and fowls) also is reported. •Kalat, J.W. 1974. Taste salience depends on novelty, not concentration, in taste-aversion learning in the rat. J. Comp. Physiol. Psycho. 86:47-50. 125 Reports the results of testing lab rats for taste salience, and indicates that the rats tend to form aversions to the novelty of the taste (or unfamiliarity), not on the "strength" of the taste. Kalat, J.W. and P. Rozin. 1970. "Salience": a factor which can override temporal continuity in taste-aversion learning. J. Comp. Physiol. Psychol. 71:192-197. Reports the evaluations of the relative importance of temporal conti nuity and specific properties of novel test solutions in establishing associations with poisons by female white-rats. Kalat, J.W. and P. Rozin. 1973. Learned safety as a mechanism in long-delay taste-aversion learning in rats, J. Comp.- Physiol. Psychol. 83:198-207. Reports the test of lab rats with lithium chloride for taste-aversion, indicates that the rats learn during the long CS-US delay that the ingested solution is "safe." 'Kapoor, I.P. and R.N. Khare. 1965. Studies on the bird and rodent control in rice mills. Bull. Grain Technol. 3:51-55. Discusses the loss to grain storage done by rodents and birds. Zn3P2 is a poison used for rodent control at 5% level by weight in whole "pakaras" (made of gram and vegetable). Kaukeinen, D.E. 1978. Methods for censusing rodents in rodenticide evalu ations. ASTM Symposium on Test Methods for Vertebrate Pest Control, Sacramento, California, March 10 (in press). ■ Describes census techniques using bait consumption and tracking board for evaluating the efficacy of a rodenticide in field conditions. Use of snap traps immediately following the post-treatment census 126 is suggested. Khan, J.A. 1974. Laboratory ejq>eriments on the food preferences of the black rat (Rattus rattus L.). Zool. J. Linn. Soc. 54:167-184. Food preferences of the black rat are described. Reports the results of testing colonies of black rats with different foods (two kinds at a time); indicates 1) of whole cereals, millet was preferred to wheat and maize, 2) semolina flour to millet flour, maize flour, and whole meal, 3) rats showed preference for moist foods and for food with groundnut oil or cane-sugar, and 4) groundnut oil was preferred to sugar. Krishnakumari, M.K. 1973. Effects of early food experience on later food preferences in adult rats. Pest Control (41):36, 38, 43. Reports the results of study of the effect of early food ejq>eriences in Norway rats (Rattus norvegicus), indicates that early experiences have little influence on food preferences of adult Norway rats. Krishnakumurthy, K. and P. Singh. 1967. Studies on rodents and their control. Part II. Control of field rats with aluminum phosphide. Bull. Grain Techol. 5(3):173-175. Reports the use of acid clay (20% sulphuric acid) impregnated with Zn3P2 for rat burrows. Percentage of success in field trials varied from place to place due to different moisture content of the soils; however, the results were not satisfactory. Kusano, T. 1977. Practical efficiency of several rodenticides on the Japanese field voles, Microtus montebelli, in agricultural areas. J. Fac. Agric., Tottori Univ. 12:8-19. Explains the use of various rodenticides (i.e, thallium sulfate, Zn3P2, 1080, norbormide, thallium acetate, thiosemicarbazide, scilliroside) 127 for controlling Japanese field voles, Microtus montebelli, in agri cultural areas. Efficiency of each rodenticide is reported also. Lord, R.D., A.M. Vilches, J.I. Maiztegui, and C.A. Soldini. 1970. The tracking board: a relative census technique for studying rodents. J. Mammal. 51(4):828-829. Describes a census technique using the tracking board for evaluating the relative abundance of rodents in different habitats in Argentina. A system of estimating the number of tracks on each board is suggested for future study. Ludeman, J.A. 1962. Control of meadow mice, kangaroo rats, prairie dogs, and cotton rats. Proc. 1st Vert. Pest Control Conf., Sacramento, California:144-163. Briefly discusses ecology of meadow mice (genus Microtus), kangaroo rats (genus Dipodomys), prairie dogs (genus Cynomys), and cotton rats (genus Sigmodon); suggests the use of acute rodenticides (i.e., zinc phosphide, strychnine, thallium sulphate, and 1080) for con trolling them. . Marsh, R.E. 1966. Methods of controlling rodents and birds in rice fields. Congres de la Protection des Cultures Tropicales, Chambre de Commerce et d’Industrie, Marseille:633-637, 998 p. Discusses damages to rice by rodents [i.e., rats genus Rattus, Muskrat (Ondatra zibethica), and nutria (Myocastor coypus), and birds, (e.g., waterfowl, red-wing blackbirds, etc.]; control methods for these vertebrate pests are suggested. Zinc phosphide grain baits at 1% or 2% are suggested for controlling rats in the rice fields. 128 Other rodenticides (i.e., compound 1080, warfarin, Pival, Fumarin, diphacinone, and PMP) are recommended for use in controlling rats in rice fields. Marsh, R.E. 1973. Recent developments in tracking dust. Proc. Rodent Control Conf. New York State Dept. of Health, Glens Falls, NY: 60-62. Reports the results of testing house mice (Mus musculus) with zinc phosphide tracking powder; suggests that it is possible to use 10 or 5% zinc phosphide tracking powder on operational basis. Marsh, R.E., W.E. Howard, and S.D. Palmateer. 1970. Effects of odors of rodenticides and adherents on attractiveness of oats to ground squirrels. J. Wildl. Manage. 34(4):821-825. Reports the results of testing ground squirrels (Spermophilus beech- eyi douglassii) with the odors of rodenticides (i.e., strychnine, sodium fluoroacetate, zinc phosphide) and of adherents (i.e., lecithin-mineral oil, starch paste, Rhoplex AC-33, and Dow latex 512-R). It was found that odor of zinc phosphide was attractive to ground squirrel. Melnick, D. 1950. Monosodium glutamate-improver of natural food flavors. Scient. Monthly 70:199-204. The introduction of monosodium glutamate into foods for human con sumption, its method of production, the mechanism by which it enhances food acceptability, and its physiological functions in the mammalian organism are discussed. Mohana Rao, A.M.K. and B.S. Rajabai. 1978. Bait shyness in two species 129 of Mus. Unpublished paper, 7 pp. The results of study of bait shyness development to zinc phosphide and its persistence in the Indian field mouse (Mus booduga Gray) and the spiny field mouse (Mus platytrix Bennett) are reported. Both species of Mus developed bait shyness and aversion to zinc phosphide after one day's exposure to sublethal dose. The aversion persisted for 95 days in former species and 75 days in latter species. Nachman, M. and J.H. Ashe. 1973. Learned taste aversions in rats as a function of dosage, concentration, and route of administrations of LiCl. Physiology and Behavior 10:73-78. Reports the results of testing male lab rats with LiCl. The rats were intraperitoneally injected with various volumes of 0.15 M of LiCl to produce a learned taste aversion to the sucrose solution. The threshold dose is discussed in relation to the amount normally given to human patients as a therapeutic dose. Nachman, M. and P.L. Hartley. 1975. Role of illness in producing learned taste aversions in rats: a comparison of several rodenticides. J. Comp. Physiol, and Psychol. 89(9):1010-1018. Reports the effects of various rodenticides (i.e., lithium chloride, warfarin, copper sulfate, sodium fluoroacetate, thallium sulfate, sodium cyanide, red squill, and strychnine sulfate). Aspects of aversions are discussed in terms of onset of symptoms, duration of symptoms, and kinds of physiological effects necessary to produce aversion. Nachman, M. and D.R. Jones. 1974. Learned taste aversions over long 130 delays in rats: the role of learned safety. J. Coup. Phsyiol. and Psychol. 86:949-956. ( ' ■ Reports the results of testing lab rats for learned taste aversions; indicates that rats learn that an ingested nontoxic solution is safe and that this learned safety may reduce aversion when the inges tion of a formerly safe solution is followed by illness. Oderkirk, G.C. 1961. Better baits for rats and mice. Pest Control 29 (8):9, 10, 12. Reports the results of testing wild rats for food preference; indi cates that the rats preferred the corn meal-oats mixture to a food bait containing only corn or oats. Flavor substances (e.g., anise oil, garlic powder) are found to improve acceptance of the squill bait when they are used in very small amounts. Peoples, S.A. 1970. The pharmacology of rodenticides. Proc. 4th Vert. Pest Control Conf., West Sacramento, California, 15-18. Discusses the primary pharmacological mechanism of the toxic action of various rodenticides-both anticoagulants (e.g., Dicoumarol, Coumachlor, Warfarin, Pindone, or Pival) and acute rodenticides (e.g., ANTU, 1080, strychnine sulfate, red squill, norbor^nide, zinc phosphide, thallium sulfate, barium carbonate, phosphorus yellow, and white arsenic). Toxicity of fumigants (e.g., carbon monoxide, methyl bromide, hydrogen cyanide) also is discussed. Prakash, I. and A.P. Jain. 1971. Bait shyness of two gerbils, Tatera indica Hardwicke and Meriones hurrianae Jerdon. Annals of Applied Biology 69:169-172. . 131 Reports the results of testing two species of gerbils with zinc phosphide for producing bait shyness. The use of zinc phosphide bait for controlling Norway rats in the fields also is discussed. Prakash, I. and P. Ojha. 1977. The phenomenon of poison-bait aversion and its possible attenuation in the Indian gerbil (Tatera indica indica). National programme for rodent pest management. Rodent Newsletter, Jodhpur Vol. 1:6-7. Reports the results of experiments carried out to overcome bait shyness in the Indian gerbil (Tatera indica indica}, indicates that poisoning attempts in different baiting material with zinc phosphide alone, after first exposure, do not reduce the development of bait shyness. Rana, B.D., I. Prakash, and A.P. Jain. 1975. Bait shyness and poison aversion in the hairy-footed gerbil, Gerbillus gleadowi murray. Proc. the All India Rodent Seminar, Ahmedabad, India:1-3. Explains the techniques for study of bait shyness; reports that the Hairy—footed Gerbil does associate the sickness developed due to feeding upon sublethal dose of zinc phosphide to the food in which it is mixed. However, the retention of memory of exposure to zinc phosphide lasts for only 10-15 days. Rennison, B.D. 1976. A comparative field trial, conducted without pre-treatment census baiting, of the rodenticides, zinc phosphide, thallium sulfate, and Gophacide against Rattus norvegicus. J. Hyg. Camb. 77:55-62. Discusses campaign against the rats in England using zinc phosphide • 132 at 1% and 2.5% levels, thallium sulfate at 0.1% and 0.3% levels, and Gophacide at 0.3% levels in the fields. Treatments with 2.5% zinc phosphide, 0.3% thallium sulfate or 0.3% Gophacide were equally effec tive and found to be more effective than treatments with 1% zinc phosphide or 0.1% thallium sulfate. Rennison, B.D., L.E. Hammond, and G.L. Jones. 1968. A comparative trial of norbormide and zinc phosphide against Rattus norvegicus on farms. J. Hyg. Camb. 66:147-158. ' Reports the test of norbormide and zinc phosphide in the fields for controlling Rattus norvegicus and concludes that treatments with zinc phosphide at 5.0 and 2.5% are significantly more successful than those with norbormide at 1.0 and 0.5%. Revasky, S.H. and E.W. Bedarf. 1967. Association of illness with prior ingestion of novel foods. Science 115:219-220. Reports the results of testing lab rats with radiation for producing aversion to a novel food rather than to familiar food. The rats were irradiated after having tasted both a novel food and a familiar food; It was found that when they were subsequently allowed to choose between these foods, their preference for the novel food was less than thé familiar food. Richter, C.P. 1939. Salt taste thresholds of normal and adrenalectomized rats. Endocrinology 24:367-371. Reports the results of testing normal and adrenalectomized rats for their salt-taste thresholds in a choice of salt solution and distilled water; indicates that normal rats detect salt in solutions as low as 0.55% (about 1 part salt to 2000 parts water) and begin to drink more 133 salt solution than distilled water. The adrenalectomzied rats distinguish salt solution in much smaller concentrations (about 1 part of salt to 33,000 parts of water, or 0.003%). Richter, C.P. 1950. Taste and solubility of toxic compounds in poison ing rats and man. J. Comp. Physiol. Psycho. 43:358-374. Discusses general properties of some acute rodenticides (e.g., strychnine sulphate, red squill, ANTU, thallium sulfate, zinc phosphide, compound 1080) and reports LD50 in mg/kg body weight for domestic and wild rats for these rodenticides. The results of testing for the taste threshold of these rodenticides for rats also are reported. Richter, C.P. and K.H. Campbell. 1940. Taste thresholds and taste preferences of lab rats for five common sugars. J. Nutr. 20:31-46. Reports the results of study for taste threshold and taste prefer ences of lab rats for five common sugars, indicates that the lower thresholds for maltose, glucose, sucrose, and galactose were 0.06%, 0.20%, 0.57%, and 1.6% respectively, and none for lactose. Rats show the greatest preference for maltose, with the maximum concen tration of the solution at 11%, next for glucose at 10%, and sucrose at 8%, and only slight appetite for galactose at 9%, and none for lactose. Robbins, R.J. 1977. Taste aversion learning in Peromyscus. (Diss. Abstr. p. 1086H3). Ph. D. Dissertation, Mich. State Univ., 222 p. Reports the results of study for taste aversion in rodent genus Peromyscus using lithium chloride as the illness-inducing agent; ' 134 indicates that taste averion does occur in Peromyscus maniculatus and will persist indefinitely to a flavor associated with illness if non-toxic fluid is simultaneously available. Robertson, A., J.G. Campbell, and D.N. Graves. 1945. Experimental zinc phosphide poisoning in fowls. J. Comp. Path. Therapeutics. 55:290- '300. Investigates zinc phosphide poisoning in poultry and indicates that this compound is highly toxic to the fowls. Toxicity, symptoms, lesions, and analytical findings in experimental zinc phosphide poisoning are described. Robison, W.H. and H.W. Hilton. 1971. Gas chromatography of phosphine derived from zinc phosphide in sugarcane. Agric. and Food Chem. 19(5):875-878. . Reports the investigations using gas chromatography of PH3 and Zn3P2 residues in sugarcane due to the use of grain bait containing Zn3P2 for small rodent control in Hawaii. Rozin, P. 1968. Specific aversion and neophobia resulting from vitamin deficiency or poisoning in half-wild and domestic rats. J, Comp, and Physiol. Psych. 66:82-88. Reports the results of comparison of the response of half-wild and domestic rats following lithium poisoning and thiamine deficiency; indicates that both strains tended to avoid the food eaten earlier, if the food had a novel, distinctive taste, poison, and/or vitamin deficiency. Rudd, R.L. and R.E. Geneily. 1956. Pesticides: their use and toxicity 135 in relation to wildlife. Calif. Dept. Fish and Game, Game Bull. No. 7:209 p. Suggests the use of zinc phosphide as a rodenticide for field rodent control at 0.5% to 1% in a variety of grains (e.g., cracked com, oat groats, and wheat). Toxicity and symptom of zinc phosphide poisoning also are discussed. Rzoska, J. 1953. Bait shyness, a study in rat behavior. Brit. J. Anim. Behaviour 1(4):128-135. Reports the results of testing white rats’ with arsenic, red squill, and barium carbonate for producing bait shyness; concludes that bait-shy rats reject the bait material experienced earlier not the rodenticides. Schien, M.W. and H. Orgain. 1953. A preliminary analysis of garbage as food for the Norway rat. Amer. J. Trop. Med. and Hygiene 2(6):1117- 1130. The results of study for food requirements of Norway rats in the city of Baltimore, Maryland are reported. Rats generally perfer foods that will result in weight gain and avoid foods that are not useful to them. They are somewhat repelled by highly spiced foods. Schoof, H.F. 1970. Zinc phosphide as a rodenticide. Pest Control 38(5): 38, 42-44. Comprehensively discusses various aspects of zinc phosphide (i.e., properties, toxicity, and hazards). The stability of this compound when used in poison bait also is discussed. «L 136 Scott, E.M. and E. Quint. 1946. Self-selection of diet. J. Nutri. 32:113-119. Reports the results of testing lab rats for their selections of * ■ ' food between a standard diet containing flavor (e.g., diacetyl, 3 p.p.m., oil of anise, 40 p.p.m.; monosodium glutamate, 1%) and a plain standard diet; indicates that the rats show no particular preference for any of those flavors. Seligman, M.E.P. 1970. On the generality of the laws of learning. Psycho. Review 77:406-418. Describes a learned taste aversion as a classical conditioning that, rats are prepared to associate taste with illness even over long delays of reinforcement, but are contraprepared to associate taste with footshock. Other types of learning (e.g., instrumental learn ing, discrimination learning, avoidance learning) also are discussed. Shumake, S.A. 1978. Food preference behavior in birds and mammals. ACS Symposium series (67). Flavor Chemistry of Animal Foods. R.W. Bullard (ed.):22-42. Discusses various aspects of food preference behavior in birds and mammals. The results of testing bait additives (e.g., sugar, cereal flavor, etc.) for enhancing naturally preferred food of the ricefield rats also are discussed. Shumake, S.A., R.D. Thompson, and C.J. Caudill. 1971. Taste preference behavior of laboratory versus wild Norway rats. J. Comp. Physiol. Psychol. 77(3):489-494. The results of testing lab and.wild rats (Rattus norvegicus) with 137 four different stimuli (sucrose, sodium chloride, quinine hydrochlor ide, and citric acid) treated on wheat pellets at four different concentrations (1.4 x IO-6, 1.4 x 10~5, 1.4 x IO-1*, and 1.4 x 10”3 mol/g) are reported. In the case of sucrose (sweet taste), the preference for treated pellets increases significantly in direct proportion to the concentrations for both strains. Both lab and wild rats show progressive aversion to the higher concentrations of sodium chloride (salty taste), quinine hydrochloride (bitter taste), and citric acid (sour taste). However, there are no significant differences between the taste preference responses of both strains. Shuyler, H.R. 1954. The development of baits for Rattus norvegicus with special preference to initial acceptability. Ph. D. Thesis, Purdue Univ., Lafayette, Indiana, 560 p. Reports the results of study of food preference of wild rats on different varieties of food (e.g., grains, liquids, fruits, fishes, vegetables, and meats). Abilities and food behavior of rats also are discussed. Simon, F.A. and L.K. Pickering. 1976. Acute yellow phosphorus poison ing. J. Am. Med. Assoc. 235(3):1343-1344. Reports three cases of acute phosphorus poisoning; suggests the safest method of preventing poisoning from this highly toxic sub stance is elimination of this product from the market. Smith, D.F. and S. Balagura. 1969. Role of oropharyngeal factors in LiCl aversion. J. Comp. Physiol. Psychol. 69s308-310. Investigates the relative contribution of oropharyngeal and gastro- . . ' . 138 intestinal factors to LiCl aversion in rats, using a single stimulus brief-exposure drinking test; indicates that oropharyngeal stimula tion is necessary for rats to learn to avoid LiCl. Srivastava, A.S. 1968. Rodent control for increased food production. Rotary Club, Kanpur, India: 152 p. Discusses the use of rodenticides (i.e., zinc phosphide, barium car bonate, red squill, ANTU, and warfarin) for rodent control. Zinc phosphide can be used at 2% in grain baits (e.g., cracked corn, corn, or steam-crushed oats) for field mouse; and at 2% in pearl-millet or sorghum, for gerbils. Procedures for campaign against rats and mice in houses and godowns are explained also. Stephenson, J.B.P. 1967. Zinc phosphide poisoning. Arch. Environ. Hlth. 15:83-88. Various cases of zinc phosphide ingestions are reported. Symptoms and treatments of zinc phosphide poisoning also «ore described. Storer, T.I. 1958. Controlling field rodents in California. Univ. of Calif. Agri. Esq?. Sta., Circular 434:20 p. Discusses the problems of field rodents (e.g., ground squirrels, tree squirrels, pocket gophers, field mice, kangaroo rats, muskrats, and rabbits) to economic loss. Ecology of ground and tree squirrels also is described. Zinc phosphide is one of the acute rodenticides that can be used for controlling them. It is used at 1% in grain baits (i.e., oat groats, whole barley, and whole oats). Storer, T.I. and E.W. Jameson. 1965, Control of field rodents in Califor nia. Div. Agri. Sci. Univ. Calif., Circular 535:50 p. 139 Discusses economic importance of field rodents (i.e., pocket gophers, ground squirrels, etc.) and diseases that can be transmitted to man; recommends use of zinc phosphide and strychnine baits for controlling them. Zinc phosphide can be used at 1% in grain baits (i.e., oat groats, whole oats, or whole barley). Strecker, R.L., W.B. Jackson, and K.R. Barbehenn. 1962, Tests of baits and poisons, p. 219-223. IN: Pacific Island Rat Ecology - Report of a Study Made on Ponape and Adjacent Islands by R.L. Strecker, J.T. Marshall, Jr., W.B. Jackson, K.R. Barbehenn, and D.H. Johnson. T.I. Storer (ed.), Bernice P. Bishop Museum, Bull. 225: Honolulu, Hawaii, 274 p. Reports the results of testing bait bases (e.g., toasted coconut, dry rice, and ground No. 3 copra) for acceptance by field rats on Ponape in a given choice in field conditions, indicates that toasted coconut is the most acceptable bait compared to others. Grated coconut meat coated with warfarin, Pival, or Fumarin-22, with the ratios of 19:1 by weight, also is tested on Polynesian and roof rats under laboratory conditions. Tevis, L. Jr. 1956. Behavior of a population of forest mice when subjec ted to poison. J. Mammal. 37(3):358-370. Reports the results of using compound 1080 for controlling white footed mice (Peromyscus maniculatus and P. truer, the major pests of Douglas fir); indicates that mice which survived a sublethal dose of this compound are conditioned to avoid untreated seed and stop eating the bait from the bait boxes in the field trials. Q 140 Tietjen, H.P. 1976. Zinc phosphide - its development as a control agent for black-tailed prarie dogs. U.S. Dept. of Interior, Fish and Wildlife Ser., Spec. Sci. Rep. Wildlife No. 195:13 p. ' ' Reports the results of development of a zinc phosphide-treated grain bait for use against black-tailed prairie dogs (Cynomys ludovicianus); suggests that 2% zinc phosphide treated oats is an effective, econo mical, and safe treatment for control of this species. Twigg, G.I. 1962. Rodent damage to sugarcane in British Guiana. World Crops 14:150-154. Reports the use of zinc phosphide baits, arsenious oxide, and warfarin for controlling a marsh rat, Holochilus sciurius berbicensis, which causes damage to sugarcane fields. Zinc phosphide bait was used in a form of pelleted baits. Losses due to rodent damage are discussed. Von Oettingen, W.F. 1952. Poisoning: a guide to clinical diagnosis and treatment. Paul B. Hoeber Inc., New York, 524 p. Explains the diagnosis of poisonings. Symptoms and treatments of the patients being poisoned are described. Wallace, P. 1976. Animal behavior: the puzzle of flavor aversion. Science 193:989-991. Comprehensively reviews the phenomenon of flavor aversion in animals (e.g., rats, coyotes, cougars, and hawks); indicates that an animal can associate a novel flavor with a subsequently induced illness (e.g., by lithium chloride, radiation). West, R.R., M.W. Fall, and J.L. Libay. 1972. Field trial of multiple . 141 baits with zinc phosphide to protect growing rice from damage by rats (Rattus rattus mindanensis). Proc. 3rd Ann. Sci. Meeting Crop Sci. Soc. of Philippines:143-147. Reports the results of using multiple baitings with zinc phosphide bait in reducing rat populations and preventing damage to growing rice. Polished rice treated with zinc phosphide at 1% was first used after the second day’s census; then 1-cm3 chunks of fresh, mature coconut dusted with zinc phosphide (1% by weight) were applied in the following week in the same area. The result of the control program was not satisfactory, probably because bait acceptance was poor. West, R.R., W.H. Robison, and A.M. Dela Paz. 1973. Weatherability of zinc phosphide treated rice baits. The Philippine Agriculturist 56(7,8)-.258-262. Reports on the analysis of the stability of zinc phosphide-treated baits exposed to rain; indicates that bait when exposed to an inch of rain for two or three days will lose about half its toxic capacity. Wilcoxon, H.C., W.B. Dragoin, and P.A. Krai. 1971. Illness induced aversions in rats and quail: relative salience of visual and gusta tory cues. Science (171):826-828. Reports the results of testing bobwhite quail and lab rats with cyclophosphamide for producing taste aversion. Both species learn in one trial to avoid flavored water when illness is induced by such drug one half hour after drinking. The visual cue is more salient than the taste cue in quail. 142 Young, P.T. 1932. Relative food preferences of the white rat. J. Comp. Psychol. 14:297-319. Reports the study of food preferences of the white rats in a free choice between two foods; indicates that the rats show preference to the food in the following order: milk, sugar, butter-fat, wheat, and flour