Isolation, characterization, and experimental infection of Streptococcus gallolyticus subspecies pasteurianus from commercial turkeys with acute septicemia: a pilot study.

Streptococcus gallolyticus (SG) is a Gram-positive cocci found as commensal gut flora in animals and humans. SG has emerged as a cause of disease in young poults between 1 and 3 wk of age. SG is associated with septicemia resulting in acute mortality with no premonitory signs in turkeys. Three SG isolates were obtained from clinical field cases of acute septicemia of commercial turkeys and used in three independent experiments. In Experiment 1, embryos were inoculated 25 d of embryogenesis with varying concentrations of SG1, SG2, or SG3. In Experiment 2, day of hatch, poults were inoculated with varying concentrations using different routes of administration of SG1, SG2, or SG3. In Experiment 3, day of hatch, poults were inoculated with only isolate SG1 using different paths. Poults were randomly selected for necropsy on d 8 and d 15 and sampled to collect spleen, heart, and liver for SG on d 21, the remaining poults were necropsied and cultured. Samples were plated on Columbia nalidixic acid and colistin agar (CNA) (40°C, 18-24 h). Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) confirmed suspect colonies. Data were analyzed using the chi-square test of independence, testing all possible combinations to determine significance (P < 0.05). Weight data were subjected to ANOVA using JMP with significance (P < 0.05). No differences were found in BW or BWG on d 0, 8, 15, or 22. Splenomegaly, focal heart necrosis, and pericarditis were observed in all groups in experiments 1 through 3. In Experiment 3, only airsacculitis was observed in a negative control in separate isolation (P > 0.05). On d 21 of Experiment 3, increased (P < 0.05) recovery of SG from spleens were observed in co-housed negative controls, as well as poults challenged by oral gavage (P > 0.05 for d 7 and d 14). These results confirm numerous previous studies indicating that SG subsp. pasteurianus is a primary infectious microorganism that causes septicemia in young poults.

Age differences in infants' attention to patterns of different complexities.

Infants 3 weeks, 8 weeks, and 14 weeks old were presented with checkerboard patterns which differed in the number of squares they contained. The older the infants, the more complex was the checkerboard that they looked at longest. This result does not appear to depend on changes in visual acuity or accommodation related to age.

Factors influencing women's quality of life in the later half of life.

BACKGROUND: Among older women in East Asia, and Taiwan in particular, there is little research on quality of life and the health care they receive to address the symptoms of menopause. This study evaluated factors which influence quality of life among post middle-age women in Taiwan. METHODS: This cross-sectional study recruited 1250 women between 43 and 77 years of age during the year 2002. The factors investigated were demographics, menstruation status, menopausal symptoms, osteoporosis status, and use of hormone replacement therapy (HRT). SF-36 was used to assess the health-related quality of life of these women. Correlation, multiple regression and path analysis were used to test for direct and indirect relationships among the variables. RESULTS: There are statistical significances between menopause symptoms and quality of life across different age groups. Path analysis shows a direct positive effect of HRT and a direct negative effect of climacteric symptoms on both physical and mental components of quality of life. Age, marital status, education and osteoporosis also have direct and indirect effects, some positive and others negative, on the components of quality of life. CONCLUSIONS: When developing programs to enhance health in post middle-age women, consideration should be given to symptom relief as well as quality of life.

Crop immune response post-Salmonella enteritidis challenge in eight commercial egg-layer strains and specific-pathogen-free White Leghorn chickens.

The crop immune response against Salmonella Enteritidis (SE) challenge in eight commercial egg-layer strains (five white-egg layer and three brown-egg layer) and specific-pathogen-free (SPF) White Leghorn (WL) hens was investigated. Pre- and post-SE challenge mucosal immune responses within the crops were evaluated. Commercial layers and SPF WL hens were orally challenged with 10(8) CFU/ml SE PT13a and SE nalR PT13, respectively. Crop lavage samples were collected at weekly intervals from day 0 (pre-challenge) to day 25-27 postinfection (PI), and bacteriological examination was performed to monitor progression of SE infection. Crop lavage samples were analyzed for SE-lipopolysaccharide (LPS)-specific IgA using enzyme-linked immunosorbent assay (ELISA). H&E-stained slides of crop sections from day 34 PI and uninfected controls were assessed for lymphoid tissue via light microscopy. Lymphoid areas were graded based on morphology, size, and cellularity using a score 0 to 5 scale. The 0 to 5 (low to high) numerical values represented progressive increases in size and cellular density of lymphoid tissue. Bacterial culture results showed the highest percentage of SE-positive crop lavage samples from all hen groups at day 5-6 PI and day 11-12 PI. A progressive decline in percentage of SE-positive crop lavage samples did occur as time PI lengthened; however, at day 25-27 PI SE persisted in crop lavage samples from SPF WL hens and three commercial white-egg layer strains. A marked increase in SE-LPS-specific IgA was measured in crop lavage samples between day 0 and day 11-12 PI for all hen groups. Crop SE-LPS-specific IgA response remained elevated above day 0 baseline for the duration of the experiment. Well-defined score 3 to 5 lymphoid tissue aggregates were observed in crop tissue sections harvested at day 34 PI. Comparison of crop sections determined a 1.2-4.0 times increase in ratio of lymphoid tissue in day 34 PI SE-challenged hens vs. uninfected control hens.

Organ weight and serum triglyceride responses of older (80 week) commercial laying hens fed an alfalfa meal molt diet.

Since one of the costs in the commercial egg industry is that of replacement pullets, commercial egg layer managers have opted to induce molt older hens in order to extend their productive life for additional egg laying cycles. Conventional molt induction involves the complete removal of feed for several days. However, this management practice can lead to deleterious physiological responses by the hen and subsequent susceptibility to infection by pathogens. Consequently less stressful molting regimens involving the feeding of low energy diets such as alfalfa have been developed. In this study, 80 week old laying hens that were deprived of feed or fed alfalfa meal during a nine day induced molt. Full fed hens were used as the control. On day 8 serum triglycerides were quantified and on day 9 hens were euthanized and the liver, spleen, heart, intestine, pancreas, ovary, and kidney were collected and weighed. Intestinal weight were highest in the non-molted hens, lower in the hens fed alfalfa, and lower still in the hens deprived of feed. Molted hens exhibited reduced weights of liver, heart, ovary, and pancreas compared to the non-molted hens. Serum triglycerides were highest in the non-molted hens, less in feed deprived hens, and the lowest in alfalfa fed hens. These results suggest that a comparable molt could be achieved with feeding alfalfa meal to 80 week hens compared to feed deprivation.

Immunological cell and serum metabolite response of 60-week-old commercial laying hens to an alfalfa meal molt diet.

The practice of induced molting involves the restriction of light, feed removal and optionally water for 5-14 days. However, there is growing concern regarding feed removal and animal welfare issues. With this in mind, alternative diets have been developed to produce similar molting effects as that of feed deprivation. Alfalfa, which largely consists of insoluble fiber, can be used as a molting diet. In this study, heterophil and lymphocyte counts, serum chemistry, and organ weight parameters were evaluated in hens that were deprived of feed or fed alfalfa during a nine day induced molt. Full-fed hens were used as the control. Blood serum parameters assessed included calcium, magnesium, glucose, total protein, ketone bodies, uric acid, and cholesterol. White blood cells were counted and categorized by cell type. On the ninth day of the trial, the hens were euthanized and the liver, spleen, heart, intestine, pancreas, ovary, oviduct, and kidney were collected and weighed. On day 8 birds molted with alfalfa or by feed deprivation had significantly higher (P<0.05) levels of ketone bodies and cholesterol and lower levels of calcium, and magnesium compared to the full-fed hens while birds molted by feed deprivation exhibited significantly lower levels of uric acid. Birds molted by both methods exhibited significant reductions in ovary, oviduct, liver and pancreas weights and increased spleen weights when compared to the non-molted hens. On days 0, 2, and 6 there were no significant differences (P>0.05) in either heterophil or lymphocyte percentages. However, heterophil percentages were higher in feed withdrawal birds than full-fed birds on day 4 but lymphocyte percentages were higher in full-fed birds compared to feed withdrawal birds. On day 8 of the induced molt lymphocyte percentages were higher from full-fed birds when compared to feed withdrawal birds but no significant differences were detectable for heterophil percentages. Based on reproductive organ weight loss and changes in serum and immunological responses of birds during molt, it appears that alfalfa meal can be an effective molt induction alternative.

Intestinal cytokine response of commercial source broiler chicks to Salmonella typhimurium infection.

Development of molecular-based immunotherapeutic strategies for controlling Salmonella Typhimurium (ST) infection in poultry requires a better understanding of intestinal and cecal cytokine responses. Accordingly, an experiment was conducted to measure changes in intestinal cytokine expression when commercial source broiler chickens were challenged with a nalidixic acid-resistant ST. Ross broiler chicks were nonchallenged with ST (control treatment) or challenged by orally giving 7.8 x 10(6) cfu at 4 d of age (STC treatment). Each treatment consisted of 4 replicate pens with 14 chicks per pen. Expression levels of proinflammatory cytokines, interferon-gamma, and antiinflammatory interleukin (IL)-10 were determined at 5 and 10 d postchallenge (PC). Intestinal flushes were also collected from each treatment at 7 d PC to estimate IgA and IgG. Results showed an upregulation in IL-1beta mRNA in STC chicks at 5 d PC. By 10 d PC, the expression of IL-1beta was further increased and accompanied by an upregulation of IL-6 and interferon-gamma mRNA, whereas IL-10 mRNA expression decreased. It was concluded that ST induced an intestinal mucosal inflammatory response in commercial source broiler chicks less than 2 wk of age.

Enhanced gross visualization of chicken Peyer's patch: novel staining technique applied to fresh tissue specimens.

The ileal Peyer's patches (Pp), secondary gut-associated lymphoid tissue of the mucosal immune system, may serve as an important site for monitoring inflammatory and immunologic responses of the host against enteric pathogens. Chicken Pp are often difficult to observe grossly, and a simple technique to enhance visualization of the Pp is lacking. Therefore, we designed a novel staining method that is quick, easy, and accurate to aid in gross identification and recovery of the chicken Pp from fresh tissue specimens. Lower alimentary tracts were harvested from White Leghorn hens and commercial broilers. The ileocecocolic region was excised intact, flushed with deionized water to remove ingesta, and a dilute eosin-Y solution was infused. After 1 min, the eosin-Y was gently extruded. Modified-crystal violet (mCV) was then injected into the gastrointestinal segment, where on the lymphoid tissue area became apparent at the serosal surface. The distal ileal Pp was visible as a pale whitish pink ovoid-focalized area with surrounding gut tissue stained light purple. The exact Pp site could be delineated at the serosal and mucosal surface by gross assessment. Light microscopy evaluation of hematoxylin and eosin-stained tissue slides prepared from the excised Pp site revealed lymphoid tissue aggregations with multiple follicular units indicative of Pp. The novel eosin-Y + mCV staining technique promotes rapid identification and accurate recovery of chicken Pp lymphoid tissue from fresh tissue specimens.

Promoting effects of polychlorinated biphenyls (Aroclor 1254) and polychlorinated dibenzofuran-free Aroclor 1254 on diethylnitrosamine-induced tumorigenesis in the rat.

The hepatic tumor-promoting activity of a commercial polychlorinated biphenyl mixture, Aroclor 1254 (AR 1254), with and without its intrinsic polychlorinated dibenzofuran (PCDF) impurities, was investigated. Male Sprague-Dawley non-inbred albino rats were treated with 66 microgram diethylnitrosamine (DENA)/ml drinking water for 5 weeks and subsequently given a control diet or a diet supplemented (100 ppm for 18 wk) with either AR 1254 or AR 1254 from which the PCDF moieties were removed (AR 1254-PCDF). Of those animals receiving DENA alone, 16% exhibited hepatocellular carcinomas. Of those rats treated with DENA followed by administration of AR 1254 or AR 1254-PCDF, 64 or 84%, respectively, developed hepatocellular carcinomas. Thus promotion with either AR 1254 or AR 1254-PCDF significantly (P less than 0.05) increased the incidence of DENA-initiated hepatocellular carcinomas. Administration of AR 1254 or AR 1254-PCDF alone did not induce hepatic tumors. Therefore, PCDF impurities were not necessary for the promoting activity of AR 1254.

The effect of feed deprivation on tissue invasion by Salmonella enteritidis.

A tissue culture procedure was utilized to compare tissue cell invasion by Salmonella enteritidis from molted and full feed hens. Three identical trials were performed in which 80-wk-old active laying hens were divided into 2 groups of 6 birds each. The molted hen group was subjected to a 14-d feed withdrawal, and the full-fed hen group was administered a standard layer ration. After feed treatment, crop, ileum, cecum, and ovary (small and large yellow follicles removed) were collected, rinsed in PBS, and placed into 50 mL of RPMI medium. The ends of intestine and crop tissues were tied to allow attachment of Salmonella only to the lumen surface. The RPMI medium containing 10(7) to 10(8) cfu of novobiocin and nalidixic acid-resistant phage type 13 Salmonella enteritidis was injected into the lumen of the intestine and crop tissues. Additionally, ovaries were incubated in 50 mL of RPMI medium containing 10(6) to 10(7) cfu of the Salmonella enteritidis. Tissues were incubated with Salmonella at 37 degrees C for 2 h, after which tissues were placed in 50 mL of fresh RPMI medium containing 500 microg/mL of gentamicin and incubated for 5 h at 37 degrees C to remove any Salmonella that had not penetrated tissues. Tissues were rinsed, stomached in 10 mL of PBS, serially diluted, and plated onto brilliant green agar containing novobiocin and nalidixic acid for Salmonella enumeration. Salmonella invasion of ovaries was reduced in tissues from molted hens in trials 1 and 2 as compared with full-fed controls (> 1.2 log reduction) but not in trial 3. Salmonella invasion of ceca from molted hens was numerically increased in trials 1 and 2 and significantly increased in trial 3 as compared with controls (> 0.8 log increase). No significant differences in Salmonella invasion were detected for crops and ileum. These data suggest that molting may affect invasion of tissues by Salmonella enteritidis.

Effect of various combinations of alfalfa and standard layer diet on susceptibility of laying hens to Salmonella enteritidis during forced molt.

Feed deprivation is commonly used by the poultry industry to induce molting and stimulate multiple egg-laying cycles. However, feed deprivation has been observed experimentally to increase susceptibility of poultry to Salmonella infections. Previous studies indicated that alfalfa was efficacious in reducing Salmonella; the present investigation was designed to evaluate the efficacy of combined alfalfa and layer diets on Salmonella colonization. Leghorn hens over 50 wk of age were divided into 12 groups of hens and placed in individual laying cages. One week prior to dietary changes, hens were put on an 8L:16D photoperiod that continued for the 9-d experiment. Hens were challenged orally with 104 cfu of Salmonella Enteritidis (SE) on d 4 of treatment and cultured for SE at the termination of the 9-d study. Two independent experiments were conducted consisting of the following treatment groups: nonfed hens, full-fed standard commercial layer diet, 100% alfalfa meal diet, a 90% alfalfa meal/10% standard commercial layer diet, and a 70% alfalfa meal/30% standard commercial layer diet. When evaluating SE colonization in the ceca (Exp. 1), a reduction (P < 0.05) was seen in the 100% alfalfa meal diet and the 70% alfalfa meal/30% standard commercial layer diet treatment groups when compared with the controls with Log10 values of 0.54, 0.44, and 2.82, respectively. Evaluation of physiological parameters showed the alfalfa treatment groups had reductions (P < 0.05) in weight loss, ovary weight, and feed consumption when compared with the full-fed standard commercial layer diet hens, and these results were comparable with the nonfed hens. In Exp. 2, all of the treatment groups had a reduction (P < 0.05) in SE colonization of the ceca when compared with the controls. There were also similar physiological reductions in weight loss, ovary weight, and feed consumption when birds were fed the alfalfa diets in Exp. 2. These data suggest that alfalfa can potentially be combined with layer ration to limit SE infection and still induce a molt comparable with feed withdrawal.

The effect of an experimental chlorate product on Salmonella recovery of turkeys when administered prior to feed and water withdrawal.

Previously, an experimental chlorate product (ECP) has been observed to reduce Escherichia coli and Salmonella infections in swine, cattle, and broilers. The following studies were performed to investigate the effects of different concentrations and durations of administering ECP on crop and ceca Salmonella typhimurium (ST) colonization of turkeys. In 2 separate trials, each conducted with 2 replicates, 15-wk-old turkey toms were challenged with 10(7) to 10(9) cfu of ST. In Experiment 1, toms were administered 0, 0.5, 1.0, 2.0, or 4.0x of ECP (a 1.0x concentration is equivalent to a 15 mM chlorate ion concentration) in the drinking water for 38 h. In Experiment 2, toms were administered a 2x concentration of ECP in the drinking water for 0, 14, 26, or 38 h prior to water withdrawal. All treatments were followed by a 10-h water withdrawal and an 8-h feed withdrawal prior to organ sampling. In Experiment 1, turkeys provided ECP had significantly (P < 0.05) lower populations and incidences of crop (>1.4 log reduction) and ceca (>0.6 log reduction) ST as compared with control birds (2.1 and 0.94 log ST average for all trials, respectively), with little or no additional benefit from administration of higher ECP concentrations. In Experiment 2, toms provided ECP had lower populations of crop (>2.2 log reduction) and ceca (>1.5 log reduction) ST when compared with controls (3.1 and 1.8 log ST, respectively). Again, there appeared to be little benefit in longer administration intervals on quantitative reduction of ST. These experiments suggest that the ECP significantly reduces Salmonella colonization in commercial turkeys when administered prior to feed and water withdrawal.

Ovarian laying hen follicular maturation and in vitro Salmonella internalization.

Transovarian transmission of paratyphoid Salmonella is well documented and occurs at a low incidence in chickens. However, the exact mechanism of follicular invasion is not well understood. The following study investigates the ability of Salmonella to invade ovarian follicles at different stages of follicular maturity in vitro. Ovarian follicles were collected from Leghorn hens and separated into three stages of maturity: (1) large yellow follicles or F follicles (LYF), (2) small yellow follicles (SYF), and (3) small white follicles (SWF). All follicles were incubated at 37 degrees C in RPMI 1640 medium. Follicles were incubated with 1 x 10(6) CFU/mL of Salmonella typhimurium and Salmonella enteritidis sensitive to gentamicin for 2 h. Samples were then removed from the bacterial culture, and placed in medium containing gentamicin sulfate for 5 h to kill any S. typhimurium or S. enteritidis, which had not invaded the follicular membrane. After the 5 h incubation, follicles were stomached in phosphate buffered saline. Serial dilutions were made of each follicle and viable S. typhimurium and S. enteritidis cells were enumerated on brilliant green agar. Two identical trials were conducted. Data suggest that Salmonella may differentially invade ovarian follicles depending on maturity of the follicle, and that SWF may be more susceptible to S. typhimurium and S. enteritidis invasion than either the SYF or the LYF.

Effects of drinking water treatment on susceptibility of laying hens to Salmonella enteritidis during forced molt.

Feed deprivation is used in the layer industry to induce molting and stimulate multiple egg-laying cycles in laying hens. Unfortunately, the stress involved increases susceptibility to Salmonella enteritidis (SE), the risk of SE-positive eggs, and incidence of SE in internal organs. Leghorn hens over 50 wk of age were divided into 4 treatment groups of 12 hens each in experiment 1 and 3 treatment groups of 12 hens in experiments 2 and 3; hens were placed in individual laying hen cages. Treatment groups were 1) nonmolted (NM) and received feed and distilled water for 9 d, 2) force molted by feed removal for 9 d and received distilled water, 3) force molted by feed removal for 9 d and received 0.5% lactic acid (LA) in distilled water. An additional group (4) in experiment 1 only was force molted by feed removal for 9 d and received 0.5% acetic acid in distilled water. Seven days before feed removal hens were exposed to an 8L:16D photoperiod, which was continued throughout the experiment. Individual hens among all treatments were challenged orally with 10(4) SE on d 4 of feed removal. When compared with the NM treatments, weight losses were significantly higher in the M treatments, regardless of water treatments. When compared with NM treatments, crop pH was significantly higher in the M treatment receiving distilled water. Crop pH was reduced to that of the NM controls by 0.5% acetic acid in the drinking water. No consistent significant changes were observed for volatile fatty acids. The number of hens positive for SE in crop and ceca after culture and the number of SE per crop and per gram of cecal contents were higher in the M treatments, when compared with the NM treatments, but there was no effect of addition of either of the acids to the drinking water. Additional research using different acid treatment regimens may provide a tool for reducing the incidence of SE in eggs and internal organs during and following molting of laying hens.

Reduction of Salmonella enterica serovar enteritidis colonization and invasion by an alfalfa diet during molt in Leghorn hens.

The standard method for molting to stimulate multiple egg-laying cycles in laying hens is feed deprivation. However, the physiological changes within hens caused by feed deprivation increase susceptibility of the hens to Salmonella enterica serovar Enteritidis (SE) infection. In an effort to develop an alternative method to induce molting without increasing susceptibility to SE, an alfalfa diet was compared with the standard molting method for the level of ovary regression and SE colonization. Hens over 50 wk of age were divided into 3 treatment groups (12 hens/group): nonmolting by normal feeding (NM), molting by feed deprivation (M), and molting by alfalfa diet (A). Individual hens on all treatments were challenged orally with 10(5) cfu of SE on the fourth day after feed changes and were analyzed for ovary weight and SE colonization or invasion in crop contents, cecal contents, liver, spleen, and ovary on the ninth day. In 3 of the 4 trials, there was a significant decrease in SE colonization of the crop between the alfalfa diet (A) and the feed deprived molt (M). In most of the 4 trials, there was a significant reduction in SE infected organs in birds fed the alfalfa diet (A) compared with birds undergoing feed deprived molt (M). Most of the trials showed no significant difference in overall SE between A and NM. Therefore, the results of this study suggest that an alfalfa diet has the potential to be used as an alternative method for forced molting, without increasing the incidence of SE in eggs and internal organs.

Readability of self-care instructional pamphlets for diabetic patients.

The readability of written instructional materials for diabetic patients is an important consideration in their use. We assessed the readability of six commonly used patient teaching guides on diabetic self-care. In addition, two pamphlets developed by our program for diabetic patients with fifth to tenth grade reading ability were also studied as controls. The difficulty of reading each pamphlet was estimated by a novel computer program which simultaneously measures readability by seven accepted formulas. The scores derived by each formula are expressed individually, in terms of grade level difficulty, and then computed to obtain a composite mean score for each text. The composite mean scores obtained are highly reliable (a = 0.95). The six commercial pamphlets were estimated to have reading difficulty levels ranging from 5.3 to 14.1 grades. A primary factor that contributed to reading difficulty was the frequent use of polysyllabic words, including technical words. Certain individual test scores, not judged to be out of range, deviated from the mean composite scores by as much as 41.2%. The two pamphlets designed by the Kentucky Diabetes Program had composite reading difficulties of 7.5 and 8.2 grade levels, respectively. These data suggest that the use of systematic readability analysis should be carried out and reported for patient instructional material. This assessment should be carried out by multiple formulas to produce a more representative index of reading difficulty than the application of a single test.

Spontaneous gestures following right hemisphere infarct.

Neurobehavioral studies of gesturing have been largely limited to left hemisphere damaged (LHD) patients. We compared spontaneous gesturing in seven right hemisphere damaged (RHD) patients, seven LHD patients, and seven normal controls (NHD) during videotaped interviews. Two judges coded symbolic, expressive, grooming, and fidgeting gestures in 120 10-sec intervals of videotape per patient. We found that RHD patients made significantly more total gestures and grooming gestures with the hand ipsilateral to their lesion than did LHD patients. Furthermore, RHD patients made more total and grooming gestures with their right hand than NHD subjects did with either hand. There were no differences in gesture production between the right and left hands of NHD patients. These results suggest that RHD produces enhanced gesturing, particularly involving grooming behavior.

Altered regulation of adrenal steroidogenesis in 2,3,7,8-tetrachlorodibenzo-p-dioxin-treated rats.

A single treatment of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) (50 micrograms/kg) produced two distinct effects on adrenal steroidogenesis in rats 13 days post-treatment. In unstressed rats, the very low corticosterone levels early in the light phase (AM) increased 4-fold relative to ad libitum-fed control (ALC) rats, but the peak level of corticosterone that is seen late in the light phase (PM) decreased up to 40% relative to ALC rats. The AM stimulation was also observed in rats pair-fed to compensate for the diminished feed intake of TCDD-treated animals, indicating that the change results from nutritional deprivation. The PM suppression, however, was not observed in pair-fed rats. In rats given a lower dose of TCDD (15 micrograms/kg), there was no AM stimulation, whereas the suppression of the PM diurnal peak of corticosterone was retained. Plasma adrenocorticotropin (ACTH) levels and adrenal size were not changed by these treatments, indicating that TCDD affects adrenal responsiveness. TCDD did not, however, have a significant effect on corticosterone secretion in rats receiving high doses of ACTH. In control animals, the availability of cholesterol to cytochrome P-450scc limits the rate of steroidogenesis. While the specific content of the cytochrome was unaffected by TCDD, cholesterol turnover by this enzyme appeared to be affected following TCDD treatment, as evidenced by small increases in the mitochondrial levels of free cholesterol, reactive cholesterol, and in the proportion of P-450scc complexed with cholesterol relative to both ad libitum- and pair-fed controls. This accumulation of mitochondrial cholesterol following TCDD treatment is consistent with an inhibition of cholesterol metabolism at cytochrome P-450scc in vivo that is removed upon isolation of the mitochondria. These TCDD-induced increases were enhanced substantially in ACTH-stimulated rats, probably because ACTH enhances cholesterol influx into the mitochondria. Normally, substrate availability is rate limiting in cholesterol side-chain cleavage, and the AM stimulation of steroidogenesis by TCDD may result from such increased cholesterol transfer. The inhibition of cholesterol side-chain cleavage resulting from TCDD treatment may, however, only become rate limiting for corticosterone synthesis when cholesterol transfer is more substantially activated, as for peak PM secretion.

Induction of drug metabolizing enzymes in polybrominated biphenyl-fed lactating rats and their pups.

Polybrominated biphenyls (PBBs) cause a mixed-type (phenobarbital- plus 3-methylcholanthrene-like) induction of liver microsomal drug metabolizing enzymes in rats. However, 2,2',4,4',5,5'-hexabromobiphenyl and 2,2',3,4,4',5,5'-heptabromobiphenyl, which together comprise less than 80% of PBBs (FireMaster), were shown to be strictly phenobarbital-type inducers. Other components (unidentified) must therefore cause the 3-methylcholanthrene-like effects. The potential for PBBs to exert effects on neonates through milk was examined. Lactating rats were fed 0, 0.1, 1.0, or 10 ppm FireMaster for the 18 days following delivery, at which time mothers and most pups were sacrificed. Pups nursing from mothers fed 10 ppm PBBs showed significant increases in liver weights and microsomal protein, and both mothers and pups had increased cytochrome P-450, aminopyrine demethylation, benzo[a]pyrene hydroxylation, and UDP-glucuronyltransferase. Pups nursing from rats fed 1.0 ppm had increases in microsomal protein, cytochrome P-450, aminopyrine demethylation, and benzo[a]pyrene hydroxylation, while their mothers were unaffected. Several pups from the 0, 0.1, and 1.0 ppm groups were maintained on their mother's diets, raised, and allowed to mate. Their pups showed much the same responses to PBBs as did the original group of pups. The effects on both generations of adult female rats were also comparable. PBBs cause a mixed-type induction in both lactating rats and their nursing pups; PBB components responsible for both aspects of this induction must be transmitted through milk. Nursing rats are approximately tenfold more sensitive to the effects of PBBs in their mother's diets than are the dams. The approximate no-effect level for microsomal induction in nursing rats is 0.1 ppm PBBs in the diet of the adult.

Studies on the microsomal metabolism and binding of polybrominated biphenyls (PBBs).

The metabolism of polybrominated biphenyls (PBBs) was studied in vitro by using rat liver microsomes in the presence of NADPH and atmospheric O2. Quantitative recoveries of all PBBs were obtained after incubations with control or 3-methylcholanthrene (MC) induced microsomes. Of the twelve major components, losses of only peaks 1 (2,4,5,2',5'-pentabromobiphenyl) and 3 (a hexabromobiphenyl) were observed following incubations with microsomes from phenobarbital (PB)- or PBBS- pretreated rats. Of seven structurally identified PBB components, only peak 1 has a bromine-free para position. Peaks 1, 2, and 5 all have two adjacent unsubstituted carbons, yet only peak 1 is metabolized. Of two dibromobiphenyl model compounds studied, the 2,2'-congener was very rapidly metabolized by PB-induced microsomes whereas its 4,4'-isomer was not. These results suggest that the presence of a free para position is required for the metabolism of brominated biphenyls. Of lesser importance appears to be the number of bromines or the availability of two adjacent unsubstituted carbons. In vivo evidence for the metabolism of peaks 1 and 3 was also provided by their drastically diminished levels in liver and milk extracts. When a 14C-PBB mixture consisting almost exclusively of peaks 4 (2,4,5,2',4',5'-hexabromobiphenyl) and 8 (2,3,4,5,2',4',5'-heptabromobiphenyl) was incubated with PB- or PBBs- induced microsomes and NADPH, only traces of radioactivity remained with the microsomes after extensive extraction. However, less radioactivity was bound to microsomes from MC pretreated or especially control rats. No radioactivity was bound to exogenous DNA included in similar microsomal incubations, regardless of the type of microsomes used. Under the same conditions, [3H]-benzo[a]pyrene metabolites were bound to DNA, and PBB-induced microsomes enhanced this binding more than six-fold.