A reproductive study of Weiser-Maples guinea pigs]

The Weiser-Maples (WE) guinea pig strain was introduced by Backshire Co., Ltd. (USA) in 1977. We have been breeding WE strain guinea pigs for skin melanization research. The WE guinea pig colony produced 1271 pups in 417 litters from May 1978 through December 1983. Breeding date are shown below. The mean litter size was 3.05, the stillborn rate was 15.2%, the weaning rate for live-born pups was 93.5% and the sex ratio was 1.01. The average age at first vaginal membrane rupture was 31.4 days at which time body weight was 290.5g. The mean length of the first 7 estrous cycles was about 17 days, with no cyclical variation in length. The mean duration of gestation was 67.9 days. Duration of pregnancy varied with litter size. There was an inverse relationship between litter size and duration of pregnancy. Most of the pups were delivered alive in mid-pregnancy with a parturition range of 56 to 76 days. The probability of pup death depends on gestational length: the lowest incidence of mortality was seen in litters born at 70 days. The mortalities were related to litter size but not to parity. There was an inverse relationship between birth weight and litter size. In WE guinea pigs, the mean weight for a litter of 1 was 120 g; for a litter of 5, the mean body weight was 58g. Male body weights were slightly heavier than female at birth and at weaning age. The mean body weights are shown below, date of birth: female 88.3g, male 93.3g, weaning age (2 weeks): female 181.1g, male 198.8g and 30 weeks: female 758.7g, male 1018.0g. These date for WE guinea pigs are comparable to those of other strains.     

Oromandibular limb hypogenesis syndromes

The oromandibular limb hypogenesis syndrome is a group of anomalies affecting the mandible, tongue, and maxilla with or without reductive limb anomalies. Their genetic origin is uncertain, and no drug-induced teratogen has been clearly identified. Although many similarities exist on both an embryologic and clinical level, distinction between these entities is appropriate. A new classification system with these principles in mind is presented. Two cases are presented of glossopalatine ankylosis with hypodactyly representing the thirteenth and fourteenth cited in the world literature. One patient presented with a fatal pulmonary hypoplasia not previously reported in association with this syndrome. Three of the 14 cases with reductive limb anomalies reported have had fatal outcomes.     

Prenatal exposure to fluconazole: an identifiable dysmorphic phenotype

BACKGROUND: Fluconazole is a triazole antifungal used to treat mycotic infections. Fluconazole is reported to act as a teratogen when used continuously at a dosage of 400-800 mg daily. Fluconazole embryopathy was previously reported in 4 cases. The common features that were also seen in the current case include multiple synostosis (including craniosynostosis and digital synostosis), congenital heart defects, skeletal anomalies, and recognizable dysmorphic facial features. CASE: We report the case of a 9-month-old male born to a 30-year-old woman following a 37-week pregnancy. The pregnancy was complicated by maternal human immunodeficiency virus (HIV) infection and multiple drug exposures, including fluconazole (400 mg/day) until the fifth month and then from 6 months to term, efavirenz, nevirapine, methadone, dapsone, pentamidine, and trimethoprim-sulfamethoxazole. At birth the infant had seizures related to neonatal abstinence syndrome and was noted to have multiple congenital anomalies. On examination at age 9 months, he had craniosynostosis secondary to coronal and lambdoidal suture closures, shallow orbital region, hypoplastic supraorbital ridges, hypertelorism, and mild ptosis. He had radioulnar synostosis and metacarpophalangeal-proximal interphalangeal symphalangism of D2-D5 bilaterally. CONCLUSIONS: The findings of cranial synostosis, multiple symphalangism, and long-bone abnormalities in our case are typical of other reported cases of fluconazole embryopathy. Our patient showed no evidence of embryopathy due to efavirenz, and he did not have the features of Antley-Bixler or other craniosynostosis syndromes. We review the literature regarding the teratogenic effects of prenatal exposure to fluconazole and provide additional evidence that prenatal fluconazole exposure has a clearly identifiable phenotype.     

Corticosteroid-induced cleft palate in short-ear mice

Corticosteroid-induced facial clefting was examined in short-ear mice (inbred strain SEA/GnJ from Jackson Laboratory). They were found to be gentle, prolific breeders (average litter size 7.3 +/- 2.6), with no increased fetal loss due to treatment. Although they have the most "resistant" H-2 haplotype (H-2d), they were found to be highly susceptible to corticosteroid-induced clefting, demonstrating that loci other than H-2 are involved in susceptibility. The short-ear locus itself is a plausible candidate, given that this gene on chromosome 9 leads to defective mesenchymal condensation, which in turn may render these mice extremely sensitive to teratogen exposure. Another gene or genes on chromosome 9 are also possibilities.     

Cardiovascular anomalies produced by nimustine hydrochloride in the rat fetus

The cardiovascular teratogenicity of nimustine hydrochloride (ACNU) was studied in rat fetuses. This drug is a nitrosourea derivative anticancer agent and produces alkylation of DNA. Pregnant Donryu rats were treated with single doses of 10, 11 or 13 mg/kg of the teratogen at various stages during gestation. Examination of the hearts was performed by microdissection after sacrificing the animals on the 20th day of gestation. The highest frequency of cardiovascular anomalies was found in the groups treated on the 8th day of gestation, but there was no difference in the rates induced by the three dosages of ACNU administered. The most common cardiovascular anomalies observed were ventricular septal defect (76.8%) and double outlet right ventricle (10.3%). A considerable number of affected fetuses (37/263) showed complex cardiac anomalies with atrioventricular (AV) malalignment and other AV valve anomalies. These anomalies include: double inlet left ventricle, straddling AV valve, atresia or stenosis of the AV valve, and dysplastic AV valve. ACNU appears to be a useful teratogenic agent for inducing complexes of cardiac anomalies which include AV malalignment.     

Experimental hyperphenylalaninemia in the pregnant guinea pig: possible phenylalanine teratogenesis and p-chlorophenylalanine embryotoxicity

Maternal hyperphenylalaninemia (HPH) due to deficient phenylalanine (Phe) hydroxylation is a recognized human teratogen associated with an increased incidence of intrauterine growth retardation, microcephaly, congenital heart disease, and mental retardation. There are no previous reports of experimental HPH during organogenesis. Sustained HPH was produced in pregnant guinea pigs by adding 3.5% Phe and 1.0% parachlorophenylalanine (pCPA), an inhibitor of Phe hydroxylase, to standard guinea pig chow. Animals consumed the supplemented test diets from gestation day 1 until killed on gestation day 17. Examination of day 17 embryos revealed that embryonic mortality was associated only with maternal pCPA administration and was independent of the degree of maternal HPH. Embryonic malformation was associated with maternal HPH as well as maternal pCPA administration. Both maternal HPH and pCPA administration were associated with embryonic growth retardation. There was no association between maternal food intake or plasma tyrosine levels and embryonic abnormality or mortality. Both Phe and tyrosine were found to be concentrated in gestation day 17 yolk sac fluid when compared to maternal plasma Phe and tyrosine. The association of embryonic malformation and maternal HPH is consistent with human data. The embryotoxicity of pCPA requires further study and highlights the necessity of appropriate controls in models of experimental HPH.     

Effect of Antibiotics on the Symptoms of Stunting Disease of Magnolia liliiflora Plants

Treatment of diseased magnolia plants with Oxytetracycline, Baytril or Tylan did not reduce the number of symptomatic plants, but promoted shoot growth, development of symptomless leaves and flower buds. The most efficient were 500 ppm Baytril, 200 ppm Tylan and 500 or 1000 ppm Oxytetracycline. Lower concentrations of Baytril and Oxytetracycline were less effective and higher concentrations of Tylan decreased the growth of magnolia shoots. All the tested antibiotic treated and untreated magnolias were shown by polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) to contain the AY (16SrI) phytoplasma and two also to contain a phytoplasma related to apple proliferation phytoplasma group (16SrX). The results indicate that Magnolia is a natural host of phytoplasmas belonging to the aster yellows and apple proliferation phytoplasma groups, and support the suggestion that phytoplasmas are the cause of magnolia stunting disease.     

Method of application of tylosin,an antibiotic for American foulbrood control,with effects on small hive beetle (Coleoptera: Nitidulidae) populations

The method of application of the antibiotic tylosin (Tylan) for control of oxytetracycline-resistant American foulbrood (Paenibacillus larvae White) was tested in honeybee (Apis mellifera L.) colonies. A powdered sugar mixture with tylosin, applied as a dust, was efficacious in eliminating American foulbrood symptoms at a rate of 200-mg Tylan per 20 g of powdered sugar, applied at weekly intervals for 3 weeks. A second method of treatment consisting of Tylan mixed with granulated sugar and vegetable shortening and applied once as a patty, at an equivalent total dose as the dust method, to diseased colonies also effectively eliminated symptoms of disease. In all colonies treated with patties, however, small hive beetle (Aethina tumida Murray) populations significantly increased, compared with the powder sugar method or untreated controls. Bee populations in patty-treated colonies also were significantly reduced, most likely the result of the invasion and proliferation of adult and larval small hive beetles. Such reduction in colony strength was not seen in dust-treated colonies. Because of the obvious damaging populations of small hive beetles, concerns about development of disease resistance, unknown risks of residues, and lack of support by regulatory agencies for the use of the patty method, the use of the dust method of tylosin is greatly favored over the patty method.     

Congenital malformation of the skeleton in Weiser-Maples guinea pigs]

Some abnormalities were observed in the occipital bone, cervical vertebrae and thoracic vertebrae of Weiser-Maples guinea pigs. In the occipital bone, the medial basilar impression was suggested to occur in 40 (32.8%) out of 122 animals. The basilar impression was classified into right, left and both side types and observed in 24, 11 and 5 animals, respectively. The basilar impression was known to be accompanied in human with some anomalies such as platybasia, Klippel-Feil syndrome, deformation of foramen magnum and so-on. These anomalies were also observed in guinea pigs. The fusion of the axis with the 3rd cervical vertebra was observed in 12 (10.5%) out of 114 animals. The deformation was sometimes observed in the temporal, interparietal, atlas and axis as well as the occipital bone. The fusion of the 7th cervical vertebra with the 1st thoracic vertebra was found in 46 (51.7%) out of 89 animals. This fusion was thought to have no relation with the basilar impression. Weiser-Maples guinea pigs are now in 19 generations of sibmating. Because these abnormalities as mentioned above are all thought to be inherited, the selective breeding will make Weiser-Maples guinea pigs suitable for the study of the basilar impression.     

Early morphological abnormalities in splotch mouse embryos and predisposition to gene- and retinoic acid-induced neural tube defects

Genetic and environmental factors contribute to an individual's neural tube defect liability. In the mouse, the gene mutation Splotch (Sp) causes a pigmentation defect in heterozygotes while homozygotes have spina bifida +/- exencephaly. Splotch homozygotes, heterozygotes, and wild-type embryos were examined for somite number, anterior neuropore closure, and posterior neuropore length. The aim was to distinguish potentially affected homozygotes early in pathogenesis and find a morphological basis for increased teratogen susceptibility in heterozygotes. Posterior neuropore closure as well as anterior neuropore closure was significantly delayed in potentially affected Sp as compared to wild-type litter embryos exceeding the incidence found in day-10-diagnosed homozygotes. Part of this excess was attributed to a transient delay in heterozygotes which in turn might predispose to retinoic acid-induced neural tube defects. This idea was supported by an outcross of Sp heterozygote males by inbred SWV females and wild-type males by SWV where a significant increase in retinoic acid-induced neural tube defects was found in Sp carrier litters.     

The use of dysmorphology in birth defects epidemiology.

Most human teratogens have been identified by the clinical recognition of characteristic patterns of congenital anomalies among children whose mothers were exposed to a particular agent during pregnancy. Although this dysmorphologic method has been valuable, it is criticized because it is not easily amenable to statistical evaluation. Conventional birth defects epidemiological studies are designed to permit rigorous statistical assessment, but such investigations usually classify congenital anomalies without adequate consideration of their known etiological heterogeneity. It is possible to combine the best aspects of these two approaches to identifying human teratogens in a "dysmorphologic case/control study." Instead of including all available cases with a given defect, only individuals having the anomaly in the context of a multiple congenital anomaly pattern without a recognizable cause would be selected for inclusion among the case group. The frequency of exposure to the putative teratogen would be determined among these selected cases and among appropriately chosen controls; conventional statistical analysis would then be performed. Although this design reduces the size of the case group compared to a conventional case/control study, the statistical power is unchanged or increased. In addition, biological plausibility is increased by concentrating upon a group of cases that is more likely to have a teratogenic cause.     

Differential dysmorphogenesis induced by microinjection of an alkylating agent into rat conceptuses cultured in vitro

A technique of microinjection of small quantities of teratogens into extraembryonic compartments or specific organ primordium of rat conceptuses of pregnancy day 11 is described. Conceptuses microinjected with 50 nl tissue culture medium developed normally for 44-45 hr when cultured in homologous rat serum, indicating that the microinjection procedure itself did not produce any deleterious effects on growth and differentiation of embryos. Microinjection of an alkylating agent, phosphoramide mustard dissolved in tissue culture medium, into the exocoelom produced anomalous embryogenesis, consisting of retarded embryonic growth, anomalies of the neural tube, and general necrosis of various organ primordia. In contrast, the embryonic development remained relatively unaffected by microinjection of identical amounts of this alkylating agent into the amniotic cavity. However, neural-tube differentiation was markedly affected when phosphoramide mustard was injected into anterior neural-tube fluid, producing anencephalic or microcephalic embryos without significant effect on postcephalic organ differentiation. The morphogenesis of the anterior limb was unaffected by local injection of the agent into somitic tissues adjacent to the presumptive limb-bud region. Therefore, it appears that differential dysmorphogenesis could be induced by microinjection of an alkylating agent into different conceptus compartments. These results indicate that even during early embryogenesis various cell types are not equally susceptible to a given teratogen, and that the differential cytotoxicity of the teratogen toward specific embryonic or extraembryonic cells and tissues may account for embryonic anomalies characteristically produced by that agent.     

Marshall J. Edwards: discoverer of maternal hyperthermia as a human teratogen

In a series of animal studies performed over a career spanning 40 years at the University of Sydney, Professor Marshall J. Edwards investigated the hypothesis that maternal hyperthermia during gestation can be teratogenic to the developing fetus. He is one of few investigators to have discovered a known human teratogen primarily through animal studies. In 1970 he earned his Ph.D. from the University of Sydney, writing a doctoral thesis entitled "A Study of Some Factors Affecting Fertility of Animals with Particular Reference to the Effects of Hyperthermia on Gestation and Prenatal Development of the Guinea-Pig." He went on to prove that hyperthermia-induced malformations in animals involve many organs and structures, particularly the central nervous system. Other defects include craniofacial anomalies, heart defects and hypodactyly, cataracts and coloboma, kyphoscoliosis, renal anomalies, dental agenesis, and abdominal wall defects. In a series of carefully planned and executed experiments, he demonstrated that the type of defect is related to the timing of the hyperthermic insult, and analyzed the underlying mechanisms. Cell death, membrane disruption, vascular disruption, and placental infarction were all implicated in causing embryonic damage. This special article reviews the scientific discoveries and personal philosophy of Marshall J. Edwards, the discoverer of maternal hyperthermia as a human teratogen.     

Animal model systems for the study of alcohol teratology

The incidence of fetal alcohol syndrome has not been declining even though alcohol has been established as a teratogen and significant efforts have been made to educate women not to abuse alcohol during pregnancy. In addition to further educational efforts, strategies to prevent or mitigate the damages of prenatal alcohol exposure are now under development. Animal models will play a significant role in the effort to develop these strategies. Because prenatal alcohol exposure causes damage by multiple mechanisms, depending on dose, pattern, and timing of exposure, and because no species of animal is the same as the human, the choice of which animal model to use is complicated. To choose the best animal model, it is necessary to consider the specific scientific question that is being addressed and which model system is best able to address the question. Animal models that are currently in use include nonhuman primates, rodents (rats, mice, guinea pigs), large animal models (pig and sheep), the chick, and simple animals, including fish, insects, and round worms. Each model system has strengths and weaknesses, depending on the question being addressed. Simple animal models are useful in exploring basic science questions that relate to molecular biology and genetics that cannot be explored in higher-order animals, whereas higher-order animal models are useful in studying complex behaviors and validating basic science findings in an animal that is more like the human. Substantial progress in this field will require the judicious use of multiple scientific approaches that use different animal model systems.     

Alberta Congenital Anomalies Surveillance System.

The Alberta Congenital Anomalies Surveillance System was started in 1966 in response to the thalidomide tragedy earlier in the decade. It was one of four provincial surveillance systems on which the federal government relied for baseline statistics of congenital anomalies. The government now collects data from six provinces and one territory. The Alberta Congenital Anomaly Surveillance System originally depended on three types of notification to the Division of Vital Statistics, Department of Health, Government of Alberta: birth notice and certificates of death and stillbirth; increased sources of ascertainment have greatly improved data quality. We present the data for 1980-86 and compare the prevalence rates of selected anomalies with the rates from three other surveillance systems. Surveillance systems do not guarantee that a new teratogen will be detected, but they are extremely valuable for testing hypotheses regarding causation. At the very least they provide baseline data with which to compare any deviation or trend. For many, if not most, congenital anomalies total prevention is not possible; however, surveillance systems can be used to measure progress in prevention.     

Optimal offspring size in a small mammal: an exception to the tradeoff invariant life-history rule

Offspring size and number were examined in a captive population of wild guinea pigs ( Cavia aperea ), and findings were compared with models of optimal offspring size for small litters. Median and modal litter size was two, regardless of maternal size or parity. Females producing their second litter tended to have litters that were larger than average. In contrast, young females that were still growing never had litters that were larger than average. Mean offspring size decreased and variation in offspring size tended to decrease with increasing litter size. Optimal offspring size models, in which offspring survival depended on the amount of resources invested, as well as litter size, predict such a trend. Little support was found for Charnov and Downhower's (1995) tradeoff invariant life-history rule that the range in offspring sizes between litters is inversely proportional to the size of the litter. Cavia aperea may be an exception to this rule because pup mass at birth did not reflect total reproductive investment, because conversion of resources into litter mass may not be linearly related to litter size and because resources were not equally partitioned among offspring within large litters. Experimental data are needed to determine the relevance of these results among mammals in general.     

Teratogenic effects of diazepam in the hamster.

Pregnant hamsters were treated with different doses of oral and intravenous diazepam during the period of organogenesis. Teratogenic effects of diazepam were observed following oral treatment on days 8 and 10 and following intravenous treatment on day 11 of gestation. Types of malformations included cleft palate, exencephaly, limb anomalies, and hemorrhage. A dose-effect relationship was not observed. Comparison with reported literature seems to indicate that diazepam may be a mild teratogen in some species.     

The effects of methylxanthines on catecholamine-stimulated and normal chick embryos.

Dose of theophylline and caffeine which do not produce aortic arch anomalies in embryonic chicks have been shown to potentiate catecholamine-induced aortic arch malformations in that experimental animal. Theophylline (2.1 X 10(-5) mole per milliliter isotonic saline solution) potentiated the effective dose of norepinephrine more than 100 times. The greatest potentiation observed with epinephrine (2.5 X) was induced by 2.6 X 10(-5) mole caffeine. This study also demonstrated that both methylxanthines specifically induce aneurysms of the ascending aorta and complete absence (or nearly complete constriction) of the right ductus arteriosus. The incidences of these types of cardiovascular malformations proved to be dose dependent with theophylline a more potent teratogen than caffeine. The mobilization of calcium and/or cyclic nucleotide phosphodiesterase inhibition by the methylxanthines are suggested as significant actions in the potentiation of catecholamine-induced aortic arch anomalies.     

Description of first germinal mosaic mutation identified in dominant skeletal mutation experiments and considerations about how to deal with this kind of spontaneous mutation in analyses

Germinal mosaicism is a well-established mechanism by which new spontaneous mutations enter the human population, but it is only rather recently that clusters of mutations arising in that way have been acknowledged and dealt with in specific-locus experiments on male mice. This paper reports the first cluster of germinal mosaic mutations to have been identified in experiments on the induction of dominant skeletal mutations. The mutation was detected in six offspring of a control male from the radiation part of an Assessment-of-Dominant-Damage (ADD) experiment. Reasons are provided to explain why this one litter of six mutants was excluded from the analysis of induction of dominant mutations causing the more common skeletal anomalies, which is reported in another paper. The effects of excluding this litter from that analysis are fully described. There is discussion of why such clusters should be included in some analyses but omitted in others. They should certainly always be reported because, in some cases, they can have a major impact on conclusions. Details on this one cluster of FCGM mutations provide numerous examples of how a dominant skeletal mutation that causes rare effects can also cause many of the more common anomalies.     

Microcomputer interfacing to mass flow type gas controllers for biotechnology applications

Summary An Apple IIe microcomputer with an Isaac analog-digital package has been interfaced to Tylan proportionating mass flow gas controllers to regulate three substrate gases which are introduced into an anaerobic bioreactor. Advantages of the system include: straightforward interfacing and programming, accurate monitoring and controlling, and advanced real time control capabilities.