Robert M. Kitchin

Induction of sister-chromatid exchanges in vivo in mice by the mycotoxins sterigmatocystin and griseofulvin

Two naturally occurring fungal mycotoxins, sterigmatocystin and griseofulvin, were tested for induction of sister-chromatid exchanges (SCEs) in bone marrow cells of female Swiss albino mice. Sterigmatocystin gave elevated SCE frequencies at all doses tested (0.06-6.0 mg/kg). In contrast, griseofulvin, tested from 0.4 to 200 mg/kg, elevated the SCE frequency only in those mice which received doses of 100 or 200 mg/kg body weight. These results indicate that both fungal mycotoxins induce SCE in vivo and are potentially mutagenic.

The effect of the breeding season, cryopreservation and physiological extender on selected sperm and semen parameters of four ferret species: implications for captive breeding in the endangered black-footed ferret

In the present investigation, comparative baseline information on selected sperm characteristics of ejaculate spermatozoa of the domestic (Mustela putorius furo), fitch (Mustela sp.) and black-footed ferrets (Mustela nigripes) and the Siberian polecat (Mustela eversmanni) are presented. The main emphasis was to establish differences and similarities among these species in relation to semen and sperm quality during the breeding season, in cryopreservation success and in supporting sperm motility in different extenders or physiological media. The results confirm that most sperm morphology abnormalities were evident during the beginning of the breeding cycle in all four species. No significant interspecies differences were apparent in the sperm attributes examined, for all sampling months during the breeding season. Moreover, all species exhibited comparable patterns of reproductive seasonality. Cryopreservation suppressed sperm characteristics equally in all species studied. Ejaculate spermatozoa of closely related ferret species shared many similar motion characteristics using computer-aided sperm motility analysis. These results suggest that the basic sperm physiology of the ferret species under examination is very similar. Disparate to the interspecies comparisons, there were significant differences for most sperm motion parameters when spermatozoa of any of the ferrets were compared in different extenders. Assisted reproductive technologies developed for use in domestic ferret, fitch ferret or Siberian polecat may be successfully applied to captive breeding of the black-footed ferret using semen during any of the functional breeding months.

Genetic Similarity of Two Forms of Cutthroat Trout, Salmo clarki, in Wyoming

Patterns of genetic differentiation among populations of cutthroat trout, Salmo clarki, in fluvial and lacustrine habitats, as well as in an area where suspected subspecies occur sympatrically were studied using protein electrophoresis. Genetic variability is very low for two genetic parameters; the proportion of loci heterozygous per individual averaged 0.01, and the proportion of loci polymorphic per population averaged 0.05. These low values are attributed to the influence of recent glaciations. The genetic identity of populations is within a narrow spectrum of 1.00 to 0.995. An estimation of the time since divergence between the two morphological forms of cutthroat trout, based on the electrophoretic identity of proteins, is 20,000 years. These results support a hypothesis of a recent origin of the fine-spotted cutthroat trout within the Snake River drainage. MERISTIC analysis demonstrates only slight differentiation among subspecies of the polytypic cutthroat trout (Salmo clarki), yet often there are obvious differences in spotting pattern and body coloration (Roscoe, 1974). However because the spotting pattern and body coloration is so variable within, as well as between major drainages the origins and evolutionary relationships among the existing subspecies are poorly understood. One such confusing group is the cutthroat trout of the Snake River above Shoshone Falls and of the

The structure―function relationships of nitrofluorenes and nitrofluorenones in the Salmonella mutagenicity and CHO sister-chromatid exchange assays

Nitrofluorenes and nitrofluorenones are bacterial mutagens and are detected in a variety of environmental pollution sources. We tested a series of nitrosubstituted fluorenes and fluorenones for their genotoxicity using both Salmonella bacteria and Chinese hamster ovary (CHO) cells to determine if structure-function relationships observed in bacteria for mutation induction are similar to those for mutations and SCE induction in mammalian (CHO) cells. The compounds studied were 2-nitrofluorene (2-NF), 2,7-dinitrofluorene (2,7-DNF), 3-nitrofluorenone (3-NFone), 2-nitrofluorenone (2-NFone), 2,7-dinitrofluorenone (2,7-DNFone), 2,4,7-trinitrofluorenone (2,4,7-TNFone), and 2,4,5,7-tetranitrofluorenone (2,4,5,7-TNFone). In bacteria, the presence of carbonyl group to convert mono-nitrofluorenes to nitrofluorenones and the addition of a second nitro group to either mono-nitrofluorene or fluorenone to form the dinitro compounds increased mutagenic activity in the Ames test. Location of the nitro group relative to the carbonyl group was important in enhancing mutagenic activity as 2-nitrofluorenone was more mutagenic than 3-nitrofluorenone. In CHO cells, the di-, tri- and tetra-nitrofluorenones were cytotoxic and delayed the progression of CHO cells through the cell cycle. The degree of the cytotoxicity could be decreased by the addition of S9. None of the compounds produced mutations when tested in the CHO/HGPRT mutation assay with the addition of S9. Nonetheless, the current study did show that these compounds, both with and without the activation by S9, can interact with the DNA and produce SCE in CHO cells. The addition of a carbonyl group had no influence on SCE frequency since both 2-nitrofluorene and 2-nitrofluorenone induced a similar frequency of SCE either with or without S9. Additional nitro groups, forming di-, tri- or tetra-nitrofluorenones, increased the frequency of SCE induced, especially when tested with S9 which limits cytotoxicity. The addition of a single nitro group to 2-nitrofluorenone did not change the SCE frequency but did cause a large increase in the frequency of mutations in bacteria. In contrast, 2,4,7-TNFone and 2,4,5,7-TNFone were less mutagenic than the 2,7-DNFone in bacteria but were more effective in production of SCE in CHO cells. This study illustrates that structure-function relationships are dependent on both the compounds tested and the type of genetic change induced.

Evaluation of the genotoxic potential of selected anti-AIDS treatment drugs at clinical doses in vivo in mice

Six anti-AIDS drugs were assessed for in vivo genotoxicity and cytotoxicity at human clinical doses with the mouse bone marrow micronucleus assay. These included four dideoxynucleosides (azidothymidine, dideoxycytidine, dideoxyadenosine, and dideoxyinosine), an anthracycline antibiotic (doxorubicin), and a chelating agent (D-penicillamine). Cytological analysis of the mouse bone marrow cells revealed: (i) The dideoxynucleosides and D-penicillamine failed to induce significant number of micronuclei, and except for one of the five doses of dideoxyinosine, none of the dideoxynucleosides were cytotoxic at the doses tested. (ii) Doxorubicin induced micronuclei in a dose-dependent manner which was statistically significant at 4-times the clinical dose but was not cytotoxic at any of the doses tested.

Intranuclear destruction of heterochromatin in two species of armored scale insects

Spermatogenesis is described in two species of armored scale insects,Parlatoria proteus andParlatoria ziziphus. In the males of both species, a haploid set of four chromosomes becomes heterochromatic during early embryogeny. The heterochromatic chromosomes are lost later by two different mechanisms during spermatogenesis. Just before meiosis begins one or more heterochromatic chromosomes disappear from each primary spermatocyte as a consequence of a rapid intranuclear chromosome destruction. Meiosis consists of a single achiasmatic division. At prophase four euchromatic and from one to three heterochromatic chromosomes are present in each cell. Although both the euchromatic and remaining heterochromatic chromosomes divide, the heterochromatic chromosomes are later eliminated by posttelophase ejection; the eliminated chromosomes then disintegrate slowly in the cytoplasm. Each of the two species displays a species specific level of heterochromatin retention and both differ in this regard from the previously describedParlatoria oleae. The evolution of a chromosome system involving intranuclear chromosome destruction is discussed.

Assessment of genotoxicity of two anti-Parkinsonian drugs (selegiline hydrochloride and bromocryptine mesylate) in vivo in mouse bone marrow cells

Selegiline hydrochloride (1-deprenyl) and bromocriptine mesylate (2-bromo-alpha-ergocryptine) are two drugs that have shown considerable promise in the treatment of Parkinsons disease. The in vivo mouse bone marrow micronucleus assay was used to examine their clastogenic and cytotoxic potential in human clinical dose range. Our results indicate that both drugs failed to induce significant number of micronuclei and were not cytotoxic at any of the doses tested, in vivo in mouse bone marrow cells, at doses as high as 16-times the clinical dose used in humans.

An in vivo giemsa chromosome banding technique

An in vivo chromosome banding technique has been developed. Swiss albino mice were injected with the DNA alkylating agents ethyl methanesulfonate, methyl methanesulfonate, or methyl ethanesulfonate 12, 24, 48 or 72 hours prior to cell harvesting. After harvesting, the cells were fixed with 3:1 methanol-acetic acid and slides were prepared by air drying. The slides were stained 2 1/2 minutes in 3% Giemsa in pH 6.8 Sorensens buffer. All three alkylating agents induced chromosome bands similar to the Giemsa bands induced by other banding techniques which involve postfixation treatments.

THE COMBINED GENOTOXIC EFFECTS OF RADIATION AND OCCUPATIONAL POLLUTANTS

Abstract The purpose of this research is to determine how short-term tests can be used as a first screen to evaluate the potential interaction between damage induced by radiation and chemicals present in the work environment. There is a potential for exposure of workers to both radiation and chemicals. Exposure standards are set one compound at a time. Little consideration is given to potential additive, antagonistic, or synergistic interactions caused by combined exposures to physical and chemical agents. Short-term tests can help identify areas of concern. We are reporting on the types of interactions observed between radiation-induced cytogenetic damage in vitro with that induced by metals, fibers, and organic solvents present in the nuclear industry and at nuclear waste sites. Synergistic interactions were observed for the induction of chromatid deletions and chromatid exchanges by combined exposure to beryllium and X rays. These aberrations are produced by radiation only during the S/G2 stages of the...