Review

Veterinary Pathology
2016, Vol. 53(5) 1057-1066
Veterinary Forensic Pathology of Animal ª The Author(s) 2016
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Sexual Abuse DOI: 10.1177/0300985816643574
vet.sagepub.com

A. W. Stern1 and M. Smith-Blackmore2

Abstract
Animal sexual abuse (ASA) involves harm inflicted on animals for the purposes of human sexual gratification and includes such
terms as bestiality, zoophilia, zoosadism, animal sexual assault, and others. The prevalence of ASA is not known, although it may be
more common than is currently perceived. Veterinarians have the skills required to identify and document cases of ASA. This
article reviews the terminology, legal definitions and forms of ASA, and its social and psychological context. An investigative
approach is outlined, including an alternate light source examination; collection of swabs for DNA analysis; sampling vaginal
washes, rectal washes, and toenails for trace evidence and biologic analyses; radiographic studies; and a complete forensic
necropsy, including histopathology. Gross lesions identified in ASA victims include injuries to the anus, rectum, penis, scrotum,
nipples, and vagina; the presence of foreign bodies; and abrasions, bruising, and other evidence of nonaccidental injury. Specialized
procedures, including examination using alternate light sources and screening tests to identify human seminal fluid within samples
from ASA victims, are of potential value but have not been validated for use in animals.

Keywords
animal sexual abuse, semen, veterinary forensic pathology, reproductive, veterinary, bestiality, interspecies sex

Animal sexual abuse (ASA) is also known by a variety of terms
(Table 1). The term animal sexual abuse is preferred in the
animal welfare field over zoophilia and bestiality.50 Bestiality,
zoophilia, and similar terms are used to explain the motives and
behavior of the perpetrator. However, as perpetrator-centric
definitions, they fail to convey a sense of the harm that occurs
to the animal. Regardless of what terminology is used, as veter-
inarians, our primary concern is the animal welfare implica-
tions of human sexual contact with animals, the recognition of
injuries, and the meticulous documentation of evidence.
ASA, like rape, is the erotization of violence, control, and
exploitation.41 ASA includes the sexual molestation of animals
by humans, including a wide range of behaviors, such as fond-
ling genitalia; vaginal, anal, or oral penetration or oral-genital
contact (from person to animal and vice versa); penetration
with an object; and injuring or killing an animal for sexual
gratification (zoosadism). The ways that ASA may result in a
fatality or injury to the animal victim differs with the activity.
ASA is an activity that may be perpetrated by men and women;
however, only men can penetrate an animal with a penis and
ejaculate on or in the animal.
Because data for various crimes are not reported in any
consistent manner, the frequency with which it occurs is
unknown. The website www.Pet-Abuse.com reports crimes
against animals (both alleged and convicted) collected from
media reports and have 108 cases currently listed since 1989
where a conviction was reached in the case or a summary
(civil) charge was issued with the abuse type ‘‘bestiality.’’ This
listing describes convictions for ASA to dogs, horses, goats,
sheep, llamas, pigs, rabbits, chickens, cows, and a guinea pig.
Veterinary pathologists should consider ASA as a possible
cause for genital trauma. While not always fatal, animals killed
in the act of ASA may be presented for necropsy. This article
provides an overview of ASA, including relevant laws with an
emphasis on the United States, as well as a description of types
of perpetrators, the forensic necropsy examination, and special
techniques for identifying and collecting trace evidence such as
semen.
Laws
Laws regarding ASA vary widely by state, and some examples
of legal definitions are shown in Supplemental Table S1. In
some states, bestiality may be a felony or a misdemeanor,
whereas in other states, it may not be prohibited. For example,
1
Veterinary Diagnostic Laboratory, University of Illinois, Urbana, IL, USA
2
Cummings School of Veterinary Medicine, Tufts University, North Grafton,
MA, USA
Supplemental material for this article is available on the Veterinary Pathology
website at http://vet.sagepub.com/supplemental.
Corresponding Author:
A. W. Stern, DVM, CMI-IV, CFC, DACVP, Veterinary Diagnostic Laboratory,
University of Illinois, 2001 S Lincoln Ave, Urbana, IL 61802, USA.
Email: awstern@illinois.edu
1058 Veterinary Pathology 53(5)

Table 1. Generally Accepted Terminology in the Social Sciences Regarding Sexual Contact With Animals.

Animal sexual abuse
(This is the preferred and encompassing
term for all sexual contact between Derived from ‘‘child sexual abuse,’’a the term used in pediatrics. Child sexual abuse is ‘‘contact or
people and animals.)
Bestiality
Opportunistic zoophile
Necrozoophile; necrozoobestialist
Contact or interaction between an animal and person, when the animal is being used for the sexual
stimulation of that person or another person.
interaction between a child and an adult, when the child is being used for the sexual stimulation of
that adult or another person.’’b Considered more precise and encompassing than either bestiality
or zoophilia.a
Used to mean sexual activity between a person and an animal.c Further understood to mean using
animals as sex objects without emotional attachment.d
A person who would be content with sexual activity with humans but will have sex with an animal if
the opportunity arises.e
A person sexually attracted to dead animals.e

Zoophile; classic zoophile Used to mean sexual activity between a person and an animal.c Further understood to mean sexual
fixation on animals; zoophiles prefer animals over people as their sexual partners.f
Zoophile by proxy An individual forced into a sex act with an animal by a human partner.e

Zoosexualist Claims an emotional as well as a perceived mutual sexual attraction to or perceived mutual
relationship with an animal.f Synonymous with zoophile.
Zooerasty Sexual contact with animals and a lack of emotional connection with an animal.g
Zoosadism An extreme form of sexual abuse that involves great suffering.c A zoosadist derives sexual pleasure
from sadistic activities with an animal such as torture. Sadism includes acts of genital mutilation.h
a
Munro HMC, Thursfield MV. ‘Battered pets’: sexual abuse. J Small Ani Pract. 2001;42:333–337.
b
Roth RA. Child Sexual Abuse: Incest, Assault and Exploitation—A Special Report. Washington, DC: US Department of Health and Human Services; 1978.
c
Munro HMC. Animal sexual abuse: a veterinary taboo? Vet J. 2006;172:195–197.
d
Beetz AM. Bestiality and zoophilia: a discussion of sexual contact with animals. In: Ascione FR, ed. The International Handbook of Animal Abuse and Cruelty: Theory,
Research, and Application. West Lafayette, IN: Purdue University Press; 2008:201–220.
e
Aggrawal A. A new classification of zoophilia. J Forensic Leg Med. 2011;18:73–78.
f
Beetz A. Bestiality/zoophilia: a scarcely investigated phenomenon between crime, paraphilia and love. J Forensic Psychol Pract. 2004;4:1–36.
g
Masters REL. Forbidden Sexual Behavior and Morality. New York, NY: Julian Publishers; 1962.
h
Dietz PE, Hazelwood RR, Warren J. The sexually sadistic criminal and his offenses. J Am Acad Psychiatry Law. 1990;18:163–178.

proof of penetration is required in Nebraska.39 Currently, 36
states have laws regarding the criminalization of ASA. Laws
typically fall into 1 of 4 categories: deviant sexual acts or
intercourse, sodomy or buggery, crimes against nature, or bes-
tiality or other animal sex prohibitions. The first 3 categories
are generally older statutes using antiquated language. As new
laws are introduced, they tend to be treated as more egregious
crimes (thus more frequently felony than misdemeanor)
(C. Edwards, personal communication, 2015).
The number of prosecutions is unknown, although prosecu-
tions for ASA are uncommonly reported. Some prosecutors
have been more inclined to bring charges under public inde-
cency, breach of the peace, indecent exposure, or cruelty to
animals, rather than under a formal bestiality statute. Prosecu-
tors often allow offenders to plead to these other charges, which
impose minimal sentences. The changing of the charge prior to
a plea makes it difficult to find confirmed cases. In addition, the
media or the court often delete any reference to ‘‘sex’’ from the
information that is released to the public, which obscures the
extent of the problem. In 2016, the Federal Bureau of Investi-
gation will collect data on animal cruelty crimes through the
National Incident Based Reporting System, including ASA as 1
of the 4 categories.
Internationally, Great Britain makes it a crime to ‘‘sexually
penetrate or to be sexually penetrated by an animal,’’ and the
statute imposes a 2-year maximum penalty. Several European
countries (Germany, Sweden, and Denmark) passed laws
banning bestiality, whereas bestiality is legal in Finland and
Romania.15,17,73
Social and Psychological Context
Human sexual contact with animals is a form of animal abuse
that has impacts for the animal victim, the abuser, and society.
The sociology, criminology, legal, and medical fields use a
variety of terms to describe human sexual contact with animals
(Table 1). Most human sexuality texts have limited discussion
devoted to zoophilia.74
In the most recent edition of the Diagnostic and Statistical
Manual of Mental Disorders (DSM-5), zoophilia is not consid-
ered a diagnosable mental health problem unless it causes dis-
tress to the person who does it. 18 This may be because
zoophiles rarely seek treatment, and not all people who commit
ASA are zoophiles, but the DSM does not consider animals as
victims (a requirement for diagnosis of a paraphilic disorder
typically requires that a human be harmed). A 10-tier classifi-
cation scheme for zoophilia has been proposed ranging from
role players to exclusive zoophiles.1
Men, women, and children may be involved in ASA activ-
ities: penetrating animals with foreign objects, committing acts
of zoosadism, or in the production of animal pornography.
People may also be penetrated by a male animal or receive oral
contact to the genitals for the purposes of sexual gratification.
In several instances, the animal caused serious injury and/or
Stern and Smith-Blackmore
death of the human involved in the sexual acts. Injuries
inflicted include penile injuries,46 rectal trauma,34 anal inju-
ries,9 and fatal blunt force trauma by a calf.16
ASA may be a crime of opportunity, such as the choice of an
animal victim by a child molester to avoid the risk of being
caught or reoffending. Offenders may have a preference for
sexual gratification through the use of animals, or they may
use animals because they are nondiscretionary and available. A
variant of bestiality is necrobestiality, which is sexual gratifi-
cation toward a dead animal.1
Although the historic cliché of a farm boy with farm animals
arises from data gathered in the Kinsey report from the late
1940s, the true frequency of sexual contact with animals by
humans is not known, and few studies have investigated this in
detail.20,27 Just as there is no single ‘‘type’’ of people who
maltreat other people, there is no single type of person who
sexually abuses animals.37 The Kinsey reports on human sex-
ual behavior indicated that 8% of males and 3.6% of postpu-
bescent women had sexual contact with animals.32,33 A survey
of college students found that both men and women acknowl-
edged having sexual contact with animals.26 An online ques-
tionnaire study identified people, including men, women, and a
single transgender, who were self-defined zoophiles.74 In a
profile of men from Pakistan with sexually transmitted infec-
tions, 2 of 465 men reported bestiality.58 A study in Brazil
identified sex with animals as a risk factor for penile cancer;
171 of 492 study participants reported to have had sex with
animals.77 When comparing polygraph results versus self-
reporting in juveniles, 26 of 32 male participants either self-
reported bestiality (confirmed with polygraph testing) or
denied bestiality with self-reporting but were considered likely
to have a history of bestiality when questioned during the poly-
graph examinations.63 The prevalence of bestiality (actual sex-
ual contact and sexual fantasy) was significantly higher in
psychiatric inpatients compared to 2 control groups.2
Similar to pedophilia, there is an imbalance of power in
ASA that makes sexual contact with an animal coercive.7 Ani-
mals are neither able to consent nor report such an episode.
Animal sexual abusers are difficult to profile because they
come from every socioeconomic group, range from teenagers
to the elderly, and have many different types of human relation-
ships. One study suggests, like some pedophiles, many self-
proclaimed zoophiles are actively engaged in high-tech jobs.74
Sexual abusers of animals are more likely to have been victims
of emotional neglect and abuse as children,5 and bestiality has
been linked to subsequent aggression against humans.27 Bes-
tiality is most commonly found among violent offenders, sex
offenders, and the sexually abused.3,19,27 This is thought to be
because sexuality and aggression become developmentally
fused when people or animals are treated as objects and are
manipulated, exploited, and controlled.44 Various studies point
to a link between bestiality and other forms of animal or human
cruelty, although there is not enough research to support any
particular hypothesis. For bestialists, generalized deviation and
sexual polymorphism as descriptors fit better than a graduation
hypothesis. Much is still unknown about the sexual abusers of
1059
Figure 1. Algorithm for forensic necropsy examination of an animal
suspected or known to be a victim of sexual abuse.
animals and their propensity to violence against people because
what is known comes from few limited studies.
A zoosadist will torture an animal to death as part of seeking
sexual gratification, and these animals can have evidence of
traumatic injuries in addition to the sexual assault.6 Strangula-
tion can be seen as a concurrent injury as in human sexual
assaults where strangulation is a common cause of death in
rape-murder cases.25,35
Graphic and violent sadistic videos (‘‘crush videos’’) depict-
ing the torture, beating, burning, and/or mutilation and death of
an animal are sold on pornographic websites. The videos are
intended to provide sexual gratification for those who are
aroused by the torturing (including crushing) of animals. Ani-
mals that do not resist may be under the influence of sedatives
or tranquilizers, or they may have been stunned from blunt-
force trauma or conditioned to accept the activity.
Forensic Necropsy Examination
An algorithm for processing suspected ASA cases is presented
in Figure 1. The veterinary pathologist has received training to
identify, describe, and interpret lesions and therefore has the
skills required to examine a possible ASA victim.49 The foren-
sic necropsy includes an internal and external examination,
including appropriate and thorough documentation and evi-
dence collection.23 Prior to performing the forensic necropsy,
a complete history should be provided to the pathologist,23
including the circumstances of the event, type of sexual contact
suspected, use of weapons/objects and/or restraints, description
of medical intervention, and information regarding the animal,
including recent urination, defecation, or ingestion of food and/
or water. It is helpful to review scene photographs and narrative
reports from the submitting law enforcement agency, if avail-
able. Data obtained from the forensic necropsy should be docu-
mented so that it can be viewed or re-created by others and
meet the standards required for legal proceedings. Documen-
tation may include a body diagram form used to record the
locations of injuries (such as contusions, abrasions) and areas
1060
that fluoresce during an alternate light source (ALS)
examination.
The collection and management of evidence is a critical
aspect of the forensic examination. It is important that the
evidence is not inadvertently destroyed during the examination,
appropriately documented, preserved in a manner that allows
investigation by other laboratories, and considered admissible
in a court of law. The examination procedure and collection of
evidence should be consistent, whether or not there are grossly
visible lesions or discernible evidence, because negative find-
ings could be useful to exonerate an alleged suspect. The
pathologist’s duty is to the court, so the examination of the
animal and collection of evidence must be objective, unbiased,
and open to multiple possibilities, rather than directed to sup-
porting one side or the other. Answering the question of what
happened to the animal is the focus and purpose of the forensic
necropsy examination, not who did it.
The steps taken during the forensic examination of an ASA
case are outlined in Figure 1. Powder-free gloves should be
worn during the handling and examination of the animal and
changed frequently to avoid cross-contamination of samples.
At the beginning of the forensic necropsy, an ALS examination
should be performed as described in the following section;
suspect areas should be documented and photographed and
swabs taken for DNA evidence.
After the ALS examination, the animal should be carefully
and thoroughly combed over butcher paper or a clean white
sheet to identify trace evidence, including foreign (human)
hairs. Foreign material of any type (dirt, stains, twigs) should
be noted and collected as evidence. It is important to take
photographs, with and without a photo scale, of any lesions
identified on the animal.23 Clippings and scrapings of the ani-
mal’s nails should be collected for potential analysis as these
may contain DNA of the perpetrator.
Evaluation for seminal fluid, including spermatozoa, is
described in detail in a separate section. Rectal and vaginal
or penile swabs should be taken for collection of DNA evi-
dence. In cases of human penetration by a dog penis, human
DNA could potentially be recovered from these samples. Two
swabs should be collected from each site to provide samples for
both the prosecution and defense, and an unused swab should
also be submitted as a negative control.
Rectal swabs should be collected prior to use of lubricants
for a clinical examination or for obtaining a rectal tempera-
ture.12 If a condom was worn, lubricants may be identified from
samples collected from the canine vagina or anus and can
provide evidence that sexual contact occurred in cases where
seminal fluid could not be identified and may be able to iden-
tify the brand of condom. The presence of condom lubricants in
a body cavity is evidence of penetration.60 The most common
lubricant (polydimethylsiloxane [PDMS], a silicone) is not
water soluble and not absorbed but can be washed away with
urination.53 Some lubricants contain particulates (cornstarch or
amorphous silica); hence, it is important to wear powder-free
gloves during the examination and evidence collection.
Nonoxynol-9 spermicide may also be detectable from swabs.8
Veterinary Pathology 53(5)
Oral swabs for DNA analysis should be obtained from both
males and female animals. Dry samples can be collected on a
cotton-tipped swab moistened with sterile saline. The swab
should be allowed to air dry and sent in paper (not plastic) to
a forensic laboratory for analysis. If damp materials are pack-
aged in a plastic bag, mold can develop and denature the DNA
evidence collected.
After collection of swabs for DNA, a vaginal wash sample
should be collected from female animals. Using a syringe and
tubing such as a plastic urinary catheter, sterile saline is infused
into the vagina, gently aspirated, and then reinfused and
reaspirated. The wash fluid is stored in a glass tube and sub-
mitted for further analysis to a forensic laboratory such as
microscopic examination for the presence and/or motility of
spermatozoa.43,57
If the rectum is not full of feces, a rectal wash sample should
be obtained from both male and female animals to collect trace
biological evidence. The procedure is similar to performing a
vaginal wash. If the rectum is full, feces should be collected
and can be analyzed for blood, gastrointestinal parasites, and/or
analysis of seminal fluid.43
After completion of the ALS examination and trace evi-
dence collection, full-body radiographs should be taken, as
described in a later section. Following this, the animal can be
shaved to examine the skin for bruising and abrasions. The skin
of the entire body should be reflected to demonstrate areas of
bruising in the subcutaneous tissues, which could have resulted
from blunt-force injury, because bruising may be difficult to
identify externally even when the hair is shaved.
Thorough examination of the gastrointestinal tract is
required, particularly the descending colon and rectum. If there
was insertion of a foreign body into the anus, perforation of the
rectum or colon can occur, and identification of the perforation
site would be warranted. Similarly, the entire reproductive tract
should be assessed in situ and then removed from the body.
Thorough examination of the reproductive tract is required
to detect perforation by a foreign body. If anal or vaginal
lesions are identified, it is important to document their location
using a clock-face description. For example, the anal sacs are
located at the 4- and 8-o’clock position, and the ventral-most
aspect of the anus would be at the 6-o’clock position. The
distance of perforations or other lesions from the anus or vulva
should be measured.
Formalin-fixed tissues should be retained. If the animal is
repeatedly a victim of ASA or has other lesions suggestive of
nonaccidental injury, histopathology may be of value to deter-
mine their age.
Macroscopic Lesions
Animal victims of sexual abuse may have a variety of injuries
to the anus/rectum and genitalia, while others have no such
injuries despite sexual contact. Vaginal and uterine lesions in
ASA victims include recurrent vaginitis, vaginal prolapse, uter-
ine tears near the cervix, vaginal stricture, cervical scarring,
uterine hemorrhage, and the presence of intrauterine,
Stern and Smith-Blackmore 1061

intracervical, or vaginal foreign objects.29,30,48,50 Insertion and

manipulation of an unknown object within both the rectum and
vagina of five 3-month-old calves resulted in massive internal
bleeding.29 In dogs, recurrent vaginitis and vaginal bleeding
have been reported.43 Vaginal foreign objects have been recov-
ered from dogs, including a candle, stick, and a broom handle
(Fig. 2).48
In horses, vaginal evisceration of the small intestine and
colon has rarely been reported.70 In 1 report, 2 of 3 cases
resulted from perforation of the vaginal fornix dorsal to the
cervix by the penis of a stallion; however, penetration with a
foreign object may present in a similar fashion, and historical
information may prove beneficial in determining the cause of
the injury.
Anal, rectal, and perineal injuries include insertion of for-
eign objects, stab wounds in the perineal region, colonic per-
foration, anal dilation, and anal tears.43,48 Peritonitis, proctitis,
rectal fistulas, and/or rectal prolapse may occur as a sequela to
anal and colonic injuries.43 A broom stick inserted into the
rectum of an intact female dog penetrated to the level of the
liver and resulted in death.48 Dilation of the anus has been
reported in sexually abused animals, but it should be noted that
the anus may dilate after death, and this finding should not be
confused with an antemortem lesion.11,48
Injuries to the scrotum have been reported in dogs.48 Band
castration is not accepted for castration of dogs, although it is
an acceptable practice for bull calves,4 and any such evidence
should be documented and retained (Fig. 3). Traumatic castra-
tion may result in a large open wound with profuse hemor-
rhage.48 Injuries to the penis of a male animal could occur in
some situations of ASA.
Other injuries may be associated with the sexual abuse act
but are not sexual in origin.43,48 These injuries include bruising,
abrasions from fingernail scrapings, fractures, thickening of the
skin of the pinna or cartilage of the otic canal, and injuries to
internal organs such as lacerations of viscera from blunt-force
trauma, sharp-force injuries, and strangulation.11 Ocular exam-
ination may identify traumatic anterior uveitis, hyphema,
scleral injection, retinal hemorrhage, or conjunctival petechial
hemorrhage in cases of blunt-force trauma or asphyxiation.

Microscopic Lesions
There is scant scientific literature on the microscopic findings
in sexually abused animals. A complete set of tissue samples
should be collected for histopathology in all forensic necrop-
sies as the findings may support or further characterize the Figure 2. Animal sexual abuse, dog. Foreign body in the uterus of an
gross findings.23 For example, if there is bruising, histopatho- intact female mixed-breed dog (consistent with the outer casing of a
logic examination may determine the age of the lesion and vibrating personal massager). Figure 3. Infarction, scrotum, dog. A
rubber band around the left testis and scrotum resulted in infarction of
differentiate hemorrhage from congestion. Proctitis may result
the tissue. Figure 4. Florescence of foreign material on the fur of a
from the trauma of retained foreign bodies or from rectal pene- dog when viewed with a Wood’s lamp. The fluorescent material was
tration and may be microscopically evident.43 sampled, and microscopic examination revealed the presence of
Histopathologic examination could also refute the gross human spermatozoa.
findings or identify an alternative cause for gross lesions such
as acute necrotizing colitis. Since natural disease might cause
similar gross findings to those occurring in sexually abused
1062
animals, it is imperative that a histopathologic examination be
performed.
Special Techniques
Special techniques may be useful in the investigation of ASA
cases, including ALS examination, screening tests to identify
semen or seminal fluid, and diagnostic imaging. Screening tests
may raise the index of suspicion for ASA, but confirmatory test-
ing will most appropriately be done at the jurisdictional crime
laboratory. Much of the available information comes from the
human forensic literature with anecdotal uses in veterinary med-
icine; therefore, many of these techniques require further valida-
tion for use in veterinary forensic pathology. The first priority for
the evidentiary material gathered is to make it available to the
judicial process. Both the prosecution and the defense will want
the opportunity to have materials tested, so any screening tests
conducted by the veterinary pathologist should not consume the
entire available sample. If there is scant material available, it
should be preserved and forwarded to the authorities.
Alternate Light Source Examination
Alternate light sources, including lasers and high-intensity fil-
tered lights, have been adapted to medicolegal investigations.36
Nonlaser light sources can enhance the contrast of biologic
evidence against the background or cause the evidence to
fluoresce.59
Semen can be identified with an ALS as it absorbs light at
particular wavelengths and then reemits the absorbed energy at
a longer wavelength.36 The excitation spectrum of human
semen is 300 to 480 nm, and the emission spectrum is 400 to
700 nm.68 The fluorescence of human semen is best detected
using an excitation wavelength of 420 to 450 nm and viewed
through orange goggles.52,71 To clearly observe the photolumi-
nescence effect of semen, the investigator must use appropriate
filters or goggles to filter out strong excitation wavelengths.68
A Wood’s lamp is a source of ultraviolet light and emits
wavelengths of 320 to 400 nm. Wood’s lamps have been
reported to fluoresce biologic substances, including urine and
semen,22,61 but the accuracy is questionable. Wood’s lamp
evaluation of human victims of sexual assault was performed
at many hospitals with Sexual Assault Nurse Examiner pro-
grams,13 although now there is a recommendation for elimina-
tion of the Wood’s lamp during sexual assault examinations.21
In 1 study, none of the 29 semen samples fluoresced when
scanned with a Wood’s lamp,61 and another study found 0%
sensitivity for detection of semen with a Wood’s lamp.52
Other ALS are more useful for detection of semen. The
Bluemaxx BM500 (Sirchie, Medford, NJ) was 100% sensitive
for human semen, although not specific, and participants could
differentiate semen from other stains 83% of the time.52 The
Polilight, which emits wave lengths of 415 to 650 nm, had a
high frequency of false positives when examining exhibits for
seminal stains; it was suggested that this light be used as a
Veterinary Pathology 53(5)
Figure 5. Animal sexual abuse, dog. Gas lucency in the uterus of the
dog in Figure 2. Radiograph.
screening aid to locate stains and that further laboratory anal-
ysis should be performed on samples of interest.71
Depending on the nature of the assault and the size of the
animal, visible internal or external wounds may or may not be
seen. In these cases, biological material found on the animal
through the use of an ALS examination may be the only marker
of animal sexual assault. The ALS examination should cover
the entire body (including the perineal region, front and back
legs, tail, ventrum, lateral body, dorsum, neck, and head). Ani-
mal victims will often self-groom the perineal region, remov-
ing trace or biological evidence, but may not be able to reach or
may be less fastidious in other areas.
There is scant literature on ALS examination of animals for
identification of seminal fluid stains. A conference proceedings
stated that a Wood’s lamp is not effective, whereas another
stated that bodily fluids will fluoresce.40,75 Examination of the
external body with a UV light or alternate light source to detect
biological evidence on ASA victims is recommended, but there
are no recommendations on the specific emitted wavelength of
light or specific light source to be used.11,43 In the experience
of 1 author (A.W.S.), the Bluemaxx BM500 was useful for
seminal fluid detection. During a Wood’s lamp examination
of a sexually abused dog examined by 1 author (M.S.-B.), an
area of clumped fur fluoresced and the material was confirmed
to be human semen based on morphological identification of
human spermatozoa at a state crime laboratory (Fig. 4).
Radiographs
Full-body radiographs should be taken to identify musculoske-
letal injuries, gas pockets in the uterus or vagina (Fig. 5), or
evidence of peritonitis. The radiographs should be carefully
examined for signs of trauma, such as periosteal reactions from
repeated leg binding or muzzling. Other findings of nonacci-
dental injury may be identified, including rib, skull, and long
bone fractures, and these may be acute or chronic.
Tail injuries are reported in cases of ASA as manipulation of
the tail may dislocate or fracture the proximal coccygeal ver-
tebrae.43 This should be distinguished from tail tip fractures,
which are often accidental in nature.43 In the experience of 1
Stern and Smith-Blackmore
author (M.S.-B), chronic or repeated sexual abuse in an animal
may lead to laxity of the ligaments at the tail base, causing the
tail to be remarkably loose or unstable at the base.
Semen Evaluation
The survival of sperm in various environments has been docu-
mented in the human literature, and sperm survival is shorter in
deceased than living humans.56 In humans, sperm may be
recovered from the vagina for an average of 23 to 38 hours
after death and may be recovered from the rectum for 12 to 28
hours after death.56 To the authors’ knowledge, there are no
comparable studies for living or deceased animals.
When moist materials are collected (such as a moist clump
of fur), direct impressions may be made on glass slides for
microscopic identification of spermatozoa. Care must be taken
to avoid contamination when collecting or drying the samples.
Dried materials may be collected for DNA analysis by cutting
the fur and preserving it in a paper envelope (a ‘‘bindle’’ or
‘‘pharmaceutical fold’’) or by swabbing with sterile water and
cotton-tipped swabs.66,69 Alternatively, dried matter may be
collected by scraping with a scalpel blade or by using a swab
moistened with sterile saline; the latter should be air-dried
before preserving in a paper envelope. The time and anatomic
location of each sample should be recorded on an evidence
collection form. It is vital to avoid contamination by using
sterile cotton-tipped swabs and preventing contact with any
object other than the target area. An unused swab should be
submitted in a similar manner to serve as a negative control.
Swabs should be collected from the oral cavity, penis or vagina,
rectum, perineal area, and fur from regions with abnormal
staining, clumping, or abnormality observed during the ALS
examination.43
Sperm from different animal species vary in shape and size.
Sperm have 3 regions: head, midpiece, and tail. Differences in
shape and structure of the heads distinguish dog, horse, sheep,
and human sperm.10 The sperm heads are paddle-shaped in
bulls, dogs, and humans; hook-shaped in rodents; and
spindle-shaped in roosters.
Histochemical and cytochemical stains can be used to visua-
lize sperm cells. The Kernechtrot-picroindigocarmine stain
(Christmas tree stain) is most often used. Nuclear fast red dif-
ferentially stains the DNA-containing sperm head red, and the
counterstain picroindigocarmine stains the tail green-blue.
There are differences in staining patterns of sperm between
humans and dogs.64 In human spermatozoa, the acrosomal cap
stains less intensely red than the nuclear portion of the sperm
head, and the midpiece and tail stain green or blue-green. Simi-
larly in the dog, there is light staining of the acrosomal cap and
dark red staining of the postacrosomal region; however, in
contrast to human sperm heads, the dog sperm head has a third
colorless region (band) between the acrosome and postacroso-
mal region.64
With a hematoxylin-eosin stain, human sperm heads have a
very pale purple acrosomal cap and a dark purple postacroso-
mal region with a distinct line of demarcation between the
1063
regions.64 Canine sperm heads stained with hematoxylin-
eosin have a pale purple postacrosomal region and pale purple
fading to a colorless acrosomal cap.64
Collaboration with veterinary clinical pathologists and
veterinary theriogenologists for cytological evaluation smears
and washes may be helpful to screen for the presence of sperm.
However, evaluation by an accredited human forensic labora-
tory is advised to ensure the evidence is admissible in a court of
law.
Tests for Seminal Fluid
The above tests may not recover spermatozoa from some cases
of sexual abuse if the perpetrator used a condom, lacked sperm
because of testicular disease or surgical sterilization, or did not
ejaculate. In such cases, the presence of seminal fluid may be
detected by screening tests for acid phosphatase (AP) test,
prostate-specific antigen (PSA), or semenogelin (Sg) or by
mass spectrometry–based proteomics.28,31,51,65,67 The confir-
matory tests for human seminal fluid are the microscopic iden-
tification of spermatozoa, analysis of short tandem repeat loci
from the Y chromosome (Y-STRs), and mass spectrometry–
based proteomics.14,42,51,67,72 Some of the tests described in
this section are practical and easy to use, but validation studies
are lacking for their use in animals for the detection of human
seminal fluid. Some research has been published on species
cross-reactivity of several available tests.45,55,62,65,67
PSA can be used as a marker of seminal fluid.27,65,76 In
humans, PSA is produced by prostate epithelial cells and is
found in seminal fluid, prostatic fluid, male serum, and male
urine. PSA is also found in apocrine sweat glands, breast milk
of lactating women, and some breast tumors.65 Nonprostatic
fluids contain very low levels of PSA, and this should not be an
issue in sexual assault cases.24 PSA may be detected in urine of
males aged 12 to 14 years or older.62 The species cross-
reactivity of 3 different membrane test assays for human PSA
has been evaluated, using a few samples of seminal fluid from a
limited number of mammalian species, including cat, dog,
horse, pig, and bull; all samples tested negative for PSA.28
Urine from male cats and dogs showed no reaction using the
commercially available ‘‘SMITEST’’ PSA card.62 In another
study, dog semen was positive for PSA (mean 44.7 ng/ml).65
AP is primarily localized to columnar epithelial cells of the
human prostate.47 Low levels of human prostatic acid phospha-
tase messenger RNA (mRNA) expression have been reported
in nonprostatic cells of the urinary bladder, kidney, pancreas,
cervix, lung, testis, and ovary.47 AP is not influenced by the
presence or absence of spermatozoa. PSA detection is consid-
ered more reliable that AP detection after 48 hours in
humans.31 A high incidence of positive reactions obtained from
mouth swabs and a single sample from a urine stain indicates
that the presence of human saliva or urine could be misinter-
preted as an indication of possible seminal fluid.38 Seminal
fluid from pygmy chimpanzee, chimpanzee, and orangutan
tested positive with the assay for human AP, whereas samples
from mandrill, gorilla, and dog were negative.65
1064
Sg (both SgI and SgII) is secreted in human semen. Sg
assays are useful for detection of seminal plasma in forensic
samples.54,55 Semen samples from pig, bull, stallion, cat, dog,
and rooster were negative for Sg using a membrane strip
test.55,67 There was no cross-reactivity for Sg in various bodily
fluids from dogs, including semen, urine, saliva, vaginal secre-
tions, and blood, with only a single false positive detected in a
fecal sample when analyzed by a membrane strip test.67
Mass spectrometry–based proteomics can be used both as a
screening technique and as a confirmation technique for identifi-
cation of various biologic fluids. Using this technique, Sg can be
used as a biomarker to identify a biologic fluid sample as semen.72
Analysis of Y-STRs may be used to compare the DNA
profile in a fluid sample to that obtained from the suspect. One
forensic study has proposed a workflow method that has incor-
porated Y-STRs analysis if the sample in question is positive
on the screening tests such as Sg and/or PSA.42
The SPERM HY-LITER (Independent Forensics, Lombard,
IL) kit is used to specifically identify human spermatozoa and
with no cross-reactivity to spermatozoa of several other animal
species.45 The reported limitations included nonspecific back-
ground staining and an increased amount of background fluores-
cence with extreme heat fixation.45
Veterinary pathologists must understand the legal limits of
the use of these assays. Any testing of seminal fluid or sperm
should be considered a screening test. If there is insufficient
material available for both screening and confirmation
(because the sample would be consumed by the screening test),
the material should be preserved and submitted to the appro-
priate authorities.
Conclusion
ASA is likely more common than documented. The veterinary
pathologist has the necessary skills to examine cases of sus-
pected ASA. When investigating a case of potential ASA, accu-
rate documentation of lesions or lack of lesions is of crucial
importance to the forensic investigation. Documented injuries
in animals that have been sexually abused include injuries to
the anus, rectum, penis, nipples, and vagina and are directly due
to sexual contact,29,30,43,48,50 whereas other injuries are indir-
ectly related and may include abrasions (caused by fingernails)
and bruising.11,43,48 Signs of ASA can be subtle, so the exam-
ination must be meticulous. Complete documentation of the
tests and procedures performed and the results of each are
required. In any case of suspected ASA, it is recommended
that the following procedures be performed in a methodical
order to avoid both a loss of potential trace evidence and to
allow for a complete and thorough forensic examination.
 Alternate light source examination
 Collection of swabs for DNA testing
 Collection of samples for trace evidence analysis (rectal
wash, vaginal wash)
 Radiographic studies
 Complete forensic necropsy, including histopathology
Veterinary Pathology 53(5)
If present, the identification of semen and seminal fluid is of
major importance in these cases as it confirms the nature of the
crime and allows for the identification of a potential perpetrator
by law enforcement agencies through the utilization of DNA
testing. In human forensic medicine, ALS examination is a
useful tool to identify semen. Recently, it has been shown that
Wood’s lamp evaluation was not useful, whereas use of the
Bluemaxx BM500 was 100% sensitive for human semen.52,61
Alternate light sources need to be validated in veterinary for-
ensic cases to be accepted in court. Similarly, currently avail-
able screening tests are potentially useful to identify human
seminal fluid within samples of fluids and stains from ASA
victims, but these tests have not been validated for use in ani-
mals, and some differences have been observed among animal
species.45,55,62,65,67 Veterinarians and forensic investigators
should design studies to evaluate and validate the use of the
investigative tools in cases of ASA.
Declaration of Conflicting Interests
The author(s) declared no potential conflicts of interest with respect to
the research, authorship, and/or publication of this article.
Funding
The author(s) disclosed receipt of the following financial support for
the research, authorship, and/or publication of this article: Dr Smith-
Blackmore’s position is supported by a grant from the Stanton
Foundation.
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