INTERNATIONAL PROGRAMME ON CHEMICAL SAFETY
WORLD HEALTH ORGANIZATION
Toxicological evaluation of certain veterinary drug
residues in food
WHO FOOD ADDITIVES SERIES 39
Prepared by:
The forty-eighth meeting of the Joint FAO/WHO Expert
Committee on Food Additives (JECFA)
World Health Organization, Geneva 1997
DIHYDROSTREPTOMYCIN AND STREPTOMYCIN (addendum)
First draft prepared by
Dr P.L. Chamberlain
Center for Veterinary Medicine
Food and Drug Administration
Rockville, Maryland, USA
1. Explanation
2. Biological data
2.1 Toxicological studies
2.1.1 Reproductive toxicity
2.2 Observations in humans
3. Comments
4. Evaluation
5. References
1. EXPLANATION
Dihydrostreptomycin and streptomycin are aminoglycoside
antibiotics used for the treatment of bacterial infections in
food-producing animals. The two compounds were evaluated together at
the twelfth (Annex 1, reference 17) and forty-third (Annex 1,
reference 113) meetings of the Committee. The forty-third Committee
established a temporary ADI of 0-30 µg/kg bw for the combined residues
of dihydrostreptomycin and streptomycin on the basis of a NOEL for
toxicological effects of 5 mg/kg bw per day in a two-year study of
toxicity in rats treated orally. The toxicological effect was a
decrease in the body weight of males at the highest dose. A safety
factor of 200 was used. The forty-third Committee determined that
additional information to assess the potential for effects on
fertility and peri- and postnatal toxicity and information on residues
would be required for re-evaluation of dihydrostreptomycin and
streptomycin in 1997: (i) an evaluation of the results of experimental
studies on the metabolism of dihydrostreptomycin and streptomycin;
(ii) data on residues of dihydrostreptomycin and streptomycin in eggs;
and (iii) results of studies to determine the relationship between the
antimicrobial activity of the residues and their concentration, as
measured by specific chemical methods.
No new studies were conducted in laboratory animals to assess the
reproductive or peri- or postnatal toxicity of dihydrostreptomycin and
streptomycin. Instead, an expert report prepared by a recognized
expert in reproductive toxicology was provided. This report was
supplemented by published articles that were not available for review
by the forty-third Committee and field reports from veterinarians in
the United Kingdom and Ireland. None of the studies or reports met
current standards for quality assurance, but they did provide some
useful information for the evaluation of these compounds. The new data
are summarized in this monograph addendum.
2. BIOLOGICAL DATA
2.1 Toxicological studies
2.1.1 Reproductive toxicity
Pigs
An EEC Directive of 26 June 1990 (90/429/EEC) listed the health
requirements related to the import and trade of porcine semen. The
Directive required that in the absence of a negative result in an
agglutination test for Leptospira spp., boars must be treated with
two doses of streptomycin at 25 mg/kg bw given 14 days apart during a
30-day isolation period prior to their admission to an approved semen
collection centre. Adoption and implementation of this Directive would
imply that there are no concerns about the reproductive safety of
streptomycin; however, the EEC Directive did not specify the extent to
which reproductive safety was considered when the decision was made to
include this requirement.
In one field report, 18 mature boars received two injections, 14
days apart, of a combination drug containing 150 mg/ml streptomycin
and 150 mg/ml dihydrostreptomycin, as directed by the EEC Directive.
The dose administered at each injection was 25 mg/kg bw streptomycin
and dihydrostreptomycin. Treatment was administered on weeks 4 and 6,
and data were collected for nine weeks. Semen was collected once or
twice per week before and after treatment and evaluated for overall
motility, percentage of sperm showing progressive forward motility,
and average number of 90-ml insemination doses produced (at a
concentration of 28 × 106 sperm per ml).
The quality of sperm motility was decreased in comparison with
pretreatment values in two boars in week 5 and in one of these animals
in week 7. By week 8, the motility quality score of one of the boars
had returned to the average for the group, while the other remained
slightly below. Values were missing for three boars in week 5 and two
boars in week 7, and the quality of sperm motility was not evaluated
for any boar in week 9. Percentage forward motility was decreased in
week 5 for four boars, and one also showed a decrease in week 7.
Values were missing for three boars in week 5 and two in week 7. One
boar showed a decrease inweek 7 but not in week 5, and another showed
a progressive decrease in weeks 8 and 9. In week 9, the values for all
but one boar were at pretreatment levels. Decreases in the number of
semen doses per ejaculate were seen in week 5 for seven boars. Semen
from two boars was reported as 'not used' at this time; semen was 'not
used' for one during weeks 1 and 7 and for the other in weeks 5-8.
Values for week 5 were missing for three boars. The number of semen
doses was decreased in eight boars in week 7, but values at this time
were missing for two boars. In week 9, the values for all but one boar
were within the range of values observed up to and including week 4.
In week 9, semen from one boar was 'not used', and the values for two
boars were missing.
The veterinarian who submitted this field report considered that
the quality of the semen from all of these boars, with the possible
exception of one, was not adversely affected by treatment. The quality
of motility in the semen from one boar appeared to have been reduced
by treatment, and the semen from this boar was 'not used' for four
weeks. This effect may have been an incidental occurrence or
treatment-related, but the latter cannot be reliably determined from
this study owing to the lack of an untreated control group, the
absence of statistical analysis of the data, and numerous missing data
entries (Glossop, 1996).
In another field report, five young (age not stated) boars at an
artificial insemination facility were treated as required by the EEC
Directive. Records of semen collections four to six weeks later were
evaluated for volume of ejaculate, raw motility score, and
concentration. No pretreatment values and no untreated control group
were included in this study. All of the parameters evaluated were
within the normal range (Glossop, 1996).
In a field report, six boars, six to seven months of age
(approximately 100 kg bw) at an artificial insemination facility were
treated as required by the EEC Directive. Sperm morphology was
assessed in two ejaculates collected after the first injection and in
two to six ejaculates collected after the second injection.
Pretreatment morphology was also assessed. No control group was used
in this study. One boar showed a notable increase in the proportion of
abnormal sperm after treatment with streptomycin, the predominant
abnormalities being bent tails and distal protoplasmic droplets.
Another boar showed high levels of abnormal sperm before treatment
with streptomycin, and the levels increased after treatment. Such
abnormalities can result from external stress. Owing to the small
sample size and lack of an untreated control group, it could not be
determined with certainty whether the observed abnormalities were
induced by treatment or by stress (Glossop, 1996).
In another field report, 11 boars, six months of age
(approximately 100 kg bw), were given one injection of
dihydrostreptomycin at 25 mg/kg bw as routine procedure prior to entry
into a boar facility for the prevention and control of Leptospira
spp. infection. Semen from four post-treatment collections over about
one month was evaluated. Semen quality was assessed on a scale of 1 to
5 (1 being the best). Pre-injection evaluations were not available,
nor was an untreated control group included in this study. The semen
quality scores improved with time after treatment for all boars
(Wager, 1996).
In a field report, 25 sows and two gilts were each given one
injection of 15 mg/kg bw of a streptomycin/dihydrostreptomycin
combination drug before breeding. The treatment was intended to remedy
a repeated breeding problem thought to be caused by Leptospira spp.
in this herd. Service records (return rates) and litter-size records
were compared with those of a group of 25 untreated sows. The return
rate for the treated group was 33%, while that of the untreated group
was 40%. Of the nine sows in the treated group that returned to
oestrus, five were successfully bred after the second service. All 10
in the untreated group that returned to oestrus were successfully bred
after the second service; however, of these 10, six were treated with
a 15 mg/kg bw dose of streptomycin and dihydrostreptomycin before the
second service. Only one of the nine treated animals was so treated,
and this animal was successfully bred after the second service.
The records of litter size showed that the percentage born dead
of the total number of pigs born to sows and gilts that conceived at
first service was 6.8% for the treated pigs and 2.3% for the untreated
pigs. For those that conceived after the second service, the
corresponding values were 7 and 3.2%. In this herd, a value of < 7%
stillborn piglets is desirable, and action is taken when the value
exceeds 10%. The reporting veterinarian commented that although the
treated group had a higher percentage of stillborn piglets, it was
still within the normal range of variability for this parameter. The
clinical or toxicological significance of this finding is uncertain as
six of 10 animals that returned to oestrus in the untreated group were
treated before the second breeding (Wager, 1996).
Sheep
In a field report, it was stated that an injection of
streptomycin or a combination of streptomycin and dihydrostreptomycin
had been used in clinical veterinary practice for the treatment of
Leptospira hardjo infections in ewes in late pregnancy, as two doses
of 25 mg/kg bw streptomycin given 14 days apart. No apparent adverse
effects on reproduction were observed or reported in the treated
animals. The basis for this conclusion was not provided (McKeown,
1996).
Cattle
Nine bulls, approximately 18 months of age, were injected
subcutaneously with two doses of 22 mg/kg bw dihydrostreptomycin given
12 h apart. Nine additional bulls served as untreated controls. Two
samples of semen were collected 5 min apart by electroejaculation on
post-treatment days 3 and 7. The samples were evaluated for pH,
volume, percent motile spermatozoa, rate of sperm motility,
concentration of spermatozoa, and capability to withstand freezing.
The post-treatment results were compared with those of samples taken
on days 5 and 1 before treatment. The investigators noted that in view
of the time required for sperm to pass through the bovine epididymis
-- about eight days -- any treatment-related effects on the parameters
evaluated would reflect an effect of the antibiotic on epididymal
physiology, accessory sex gland function, and/or the ejaculatory
response to electrical stimulation. On each of days 8, 17, and 26
after treatment, one treated and one control bull underwent surgical
removal of the testes. Testicular size and weight were recorded, and
the samples were examined histologically at all stages of
spermatogenesis. The results showed no treatment-related effects, and
it was concluded that treatment with streptomycin had no adverse
effect on spermatogenesis in the bulls in this study (Abbitt et
al., 1984).
In a field report, the number of doses of semen obtained from an
ejaculate were recorded for four bulls used for artificial
insemination over the course of one year. After six months, each bull
received two injections of 25 mg/kg bw of a streptomycin/
dihydrostreptomycin combination drug given approximately three weeks
apart. The mean number of doses in seven ejaculates collected after
treatment increased slightly for three of the bulls, in comparison
with the mean for the seven ejaculates collected before treatment. For
the fourth bull, the mean number of doses decreased but was still
within the normal range. Treatment thus had no apparent detrimental
effect on the number of doses produced by the bulls (Kennedy, 1996).
About 1500 bulls intended for artificial insemination which were
housed at Danish centres were treated with two intramuscular doses of
25 mg/kg bw streptomycin given 14 days apart, as directed by the
Danish Bull Service Directive (88/407/EEC), which became effective in
January 1990. No clinical anomalies or deficits in semen production or
quality were reported to the Danish Veterinary Service during the year
after implementation of the Directive. As the fertility of animals in
artificial insemination centres is carefully monitored, any adverse
effects of streptomycin on semen quality would probably have been
reported (personal communication from Jorgen Bent Anderson to Kennedy,
1996).
Routine use of streptomycin or streptomycin and
dihydrostreptomycin by one veterinarian in clinical veterinary
practice for the control of infections with Leptospira spp. in about
50 dairy herds comprising 120 head per herd did not result in adverse
effects on the fertility of the treated animals. The treatment regime
described for use in pregnant cows and stock bulls included a
combination of penicillin and 250 mg/ml dihydrostreptomycin (or
streptomycin), which was injected intramuscularly at 20 ml (5 g) for
three or four consecutive days. Another regime reported was two
injections of 25 mg/kg bw 14 days apart, given as a 'blanket
treatment' over a short period of time on a given farm to all adult
breeding stock, including pregnant cows, non-pregnant cows, and stock
bulls. No adverse effects on fertility were associated with these
treatment regimes (McKeown, 1996).
Council Directive 88/407/EEC of 14 June 1988 on health
requirements for intra-community trade and imports of frozen bovine
semen stated that finally diluted semen must contain no less than 500
IU/ml streptomycin. This implies a lack of any direct adverse effects
of streptomycin on bovine semen; however, the data on which this
conclusion was based were not provided. Streptomycin in combination
with penicillin was shown to facilitate the preservation of motility
and acrosome morphology of stored boar semen by controlling bacterial
growth (Bamba & Sone, 1981).
Horses
Field reports from two veterinarians describe the use of
penicillin and streptomycin (or dihydrostreptomycin) in pregnant mares
and breeding stallions, with no obvious adverse effects on fertility
or pregnancy outcomes. The treatment regime described was 20 ml (5 g)
of the combination (containing 250 mg of streptomycin or
dihydrostreptomycin per ml) given intramuscularly once daily for five
consecutive days. Details of the examinations conducted on the
fertility of individual animals were not provided (Dillon, 1996;
Mills, undated).
2.2 Observations in humans
In a brief published article on the treatment of tuberculosis
during pregnancy, use of streptomycin was discussed. According to the
authors, streptomycin is the only licensed anti-tubercular drug with
known harmful effects on the fetus. The specific effect described was
congenital ototoxicity, attributed to eighth cranial nerve damage. No
other congenital defects due to streptomycin have been reported (Addis
& Blowey, 1996).
In a published review of the use of antibiotics for ear, nose,
and throat disorders in mothers during pregnancy or lactation, Niebyl
(1992) discussed the use of aminoglycosides. He noted that clinical
deafness has occurred in the offspring of mothers who took
streptomycin or kanamycin during pregnancy. Doses as low as 1 g
streptomycin twice weekly for eight weeks during the first trimester
caused ototoxicity in offspring. He also reported that no other
teratogenic effects are associated with the use of aminoglycosides in
the first trimester. In a study of 135 infants exposed to
streptomycin, no teratogenic effects were observed. Additional
evidence cited to support this conclusion was a study of 1619 newborns
whose mothers were treated for tuberculosis with multiple drugs,
including streptomycin, during pregnancy. The incidence of congenital
defects was the same as that in the control group. Niebyl noted that
limited information existed about the excretion of gentamicin into
breast milk, but low levels of amikacin, kanamycin, streptomycin, and
tobramycin have been detected. He concluded that ototoxicity would not
be expected to occur in infants exposed in this way in view of the low
level of excretion into breast milk and the poor oral absorption of
these drugs.
In a published review of antituberculosis therapy during
pregnancy, Scheinhorn and Angelillo (1977) addressed the use of
streptomycin by reviewing several studies, with a total of 690 cases,
in which streptomycin was administered to women during pregnancy.
Evaluation of the children born to these mothers showed 16 with loss
of high-frequency hearing, eight with vestibular defects, and one with
clinical disability. Since organogenesis of the inner ear occurs
during the seventh intrauterine week and differentiation of the
cochlear cells continues up to mid-gestation, the authors recommend
avoidance of the use of streptomycin during this period.
3. COMMENTS
The present Committee considered data from published references
and unpublished field reports on the reproductive toxicity of
dihydrostreptomycin and streptomycin. Although none of the studies or
reports met current standards for quality assurance, they did provide
some useful information for evaluation of the safety of the compounds.
The field reports consisted of empirical observations by
veterinarians and evaluations of fertility records for cattle, pigs,
sheep, and horses treated parenterally with streptomycin or a
formulation of dihydrostreptomycin and streptomycin for the treatment
or control of infections by Leptospira spp. In most cases, the
treatment consisted of two injections of 25 mg/kg bw streptomycin
given 14 days apart. Treatment regimes for cattle and horses
consisting of daily injections of 5 g of streptomycin or of the
combination for three to five consecutive days were also described.
None of these reports indicated that treatment with streptomycin or
the combination had an adverse effect on the fertility or reproductive
performance of the treated animals.
The Committee was also asked to consider European directives that
required bulls and boars to be injected with streptomycin for the
control of Leptospira spp. upon entry to artificial insemination
facilities. The treatment regimes consisted of two injections 14 days
apart of 25 mg/kg bw of streptomycin or of a combination of
dihydrohydrostreptomycin and streptomycin. In one country alone, 1500
bulls were treated over the course of one year, with no reported
adverse effects on the quality of the semen produced. The Committee
recognized that the fertility of animals in artificial insemination
centres is carefully monitored and therefore any adverse effects of
streptomycin on semen quality would probably have been reported.
In a controlled study, nine bulls were injected subcutaneously
with two doses of 22 mg/kg bw of dihydrostreptomycin 12 h apart. Semen
was collected and evaluated three and seven days after treatment, and
the results were compared with those for samples collected five and
one days before treatment. Semen samples were evaluated for pH,
volume, percentage of motile spermatozoa, rate of spermatozoal
motility, concentration of spermatozoa, and capacity to withstand
freezing. Because it takes about eight days for bovine sperm to pass
through the epididymis, these post-treatment samples represented sperm
that were in the epididymis at the time of treatment. On each of days
8, 17, and 26 after treatment, one treated and one control bull
underwent surgical removal of the testes. Testicular size and weight
were recorded, and the samples were examined histologically for all
stages of spermatogenesis. The results showed no treatment-related
effects on any of the parameters evaluated. The Committee concluded
that treatment with streptomycin had no adverse effect on
spermatogenesis in the bulls in this study.
The Committee reviewed new information on the pregnancy outcomes
of women treated parenterally with streptomycin for tuberculosis. The
new information supported the conclusion that streptomycin damages the
eighth cranial nerve in offspring born to treated mothers. Rigourous
literature searches revealed no other reported adverse reproductive
effect associated with the use of streptomycin in humans.
At the forty-third meeting, the Committee evaluated a study of
reproductive toxicity in ICR mice treated with streptomycin at 250
mg/kg bw per day intraperitoneally on days 12-18 of gestation.
Behavioural tests were conducted on the offspring. No treatment-
related adverse effects on activity or functional development were
observed. The present Committee concluded that, although the protocol
of this study allowed an evaluation of prenatal effects, the results
could not be used to evaluate the peri- or postnatal effects of
streptomycin because the dams were not treated throughout lactation.
4. EVALUATION
The Committee noted the long history of use of
dihydrostreptomycin and streptomycin in veterinary and human medicine
and the notable absence of reports in the literature of adverse
effects on fertility or peri- or postnatal toxicity other than
ototoxicity. Furthermore, the Committee recognized that very small
amounts of aminoglycosides are absorbed by adults and infants after
ingestion. It concluded that consumption of residues of
dihydrostreptomycin or streptomycin in foods derived from animals
treated in accordance with good practice in the use of veterinary
drugs presents essentially no risk to peri- or postnatal human health.
The Committee considered the potential risk for infants consuming
milk containing residues of dihydrostreptomycin or streptomycin during
the first weeks of life, before closure of the gut. The temporary MRL
for the combined residues of dihydrostreptomycin and streptomycin in
cows' milk recommended by the forty-third Committee was 100 µg/litre.
By comparison, streptomycin is administered parenterally to infants
for therapeutic purposes at doses of 10-20 mg/kg bw per day. The
present Committee concluded that the low levels of these compounds
permitted in cows' milk present essentially no risk to newborns.
The forty-third Committee determined that the ADI based on the
antimicro-bial activity of the combined residues of
dihydrostreptomycin and streptomycin is 0-80 µg/kg bw. The equation
used by the forty-third Committee was modified at the forty-seventh
meeting (Annex 1, reference 125) by replacing the value for daily
faecal bolus (150 g) with a value for colonic content (220 g). This
increases the ADI based on the antimicrobial activity of the combined
residues of dihydrostreptomycin and streptomycin to 0-120 µg/kg bw.
The forty-third Committee determined that the most sensitive
toxic effect observed in studies on dihydrostreptomycin and
streptomycin was that obtained with dihydrostreptomycin in a two-year
study of toxicity in rats treated orally. The NOEL was 5 mg/kg bw per
day on the basis of a decrease in the body-weight gain of males at the
high dose. The present Committee confirmed this as the appropriate
NOEL, applied a safety factor of 100, and established a group ADI of
0-50 µg/kg bw for the combined residues of dihydrostreptomycin and
streptomycin.
5. REFERENCES
Abbitt, B., Berndtson, W.E. & Seidel, G.E., Jr (1984) Effect of
dihydrostreptomycin or oxytetracycline on reproductive capacity of
bulls. Am. J. Vet. Res., 45, 2243-2246.
Addis, A. & Blowey, D. (1996) Tuberculosis during pregnancy. Motherisk
Newsl., 5, 4.
Bamba, K. & Sone, M. (1981) Factors affecting the quality of boar
semen stored by means of dialysis. J. Reprod. Fertil., 66, 193-197.
Dillon, H. (1996) Unpublished report from Hugh Dillon at the practice
of Edward C. Gowing, The Curragh, Kildare. Submitted to WHO by
Norbrook Laboratories Ltd, Newry, Northern Ireland, United Kingdom.
Glossop, C.E. (1996) Data on the use of streptomycin in AI boars.
Unpublished reports from Christianne E. Glossop, Merton Farmhouse, The
Street, Lea, Malmesbury, Wiltshire. Submitted to WHO by Norbrook
Laboratories Ltd, Newry, Northern Ireland, United Kingdom.
Kennedy, B.W. (1996) Review of semen collection records for four long
stay bulls for the period March 1989 to April 1990. Unpublished report
submitted by Brian W. Kennedy, Company Veterinary Surgeon, Ai Services
(Northern Ireland) Ltd, Antrim Road, Newtownabbey, Co. Antrim.
Submitted to WHO by Norbrook Laboratories Ltd, Newry, Northern
Ireland, United Kingdom.
McKeown, D.M. (1966) Unpublished report from David M. McKeown,
Knockanboy Veterinary Clinic, Dervock, Ballymoney, Antrim, Northern
Ireland. Submitted to WHO by Norbrook Laboratories Ltd, Newry,
Northern Ireland, United Kingdom.
Mills, N.J. (undated) Unpublished report from Nicholas J. Mills,
Cinque Ports Veterinary Associated, Hawkhurst, Kent. Submitted to WHO
by Norbrook Laboratories Ltd, Newry, Northern Ireland, United Kingdom.
Niebyl, J.R. (1992) Use of antibiotics for ear, nose, and throat
disorders in pregnancy and lactation. Am. J. Otolaryngol., 13,
187-192.
Scheinhorn, D.J. & Angelillo, V.A. (1977) Antituberculous therapy in
pregnancy. Risks to the fetus. West. J. Med., 127, 195-198.
Wager, A. (1996) Data on the use of dihydrostreptomycin and
streptomycin in pigs. Unpublished report from Annika Wager, Garth
Veterinary Group, Straight Lane, Beeford, Nr. Driffield, East
Yorkshire. Submitted to WHO by Norbrook Laboratories Ltd, Newry,
Northern Ireland, United Kingdom.
See Also:
Toxicological Abbreviations