Regulation of lactose, glucose and sucrose metabolisms in S. thermophilus.

Streptococcus thermophilus is a bacterium widely used in the production of yogurts and cheeses, where it efficiently ferments lactose, the saccharide naturally present in milk. It is also employed as a starter in dairy- or plant-based fermented foods that contain saccharides other than lactose (e.g., sucrose, glucose). However, little is known about how saccharide use is regulated, in particular when saccharides are mixed. Here, we determine the effect of the 5 sugars that S. thermophilus is able to use, at different concentration and when they are mixed on the promoter activities of the C-metabolism genes. Using a transcriptional fusion approach, we discovered that lactose and glucose modulated the activity of the lacS and scrA promoters in a concentration-dependent manner. When mixed with lactose, glucose also repressed the two promoter activities; when mixed with sucrose, lactose still repressed scrA promoter activity. We determined that catabolite control protein A (CcpA) played a key role in these dynamics. We also showed that promoter activity was linked with glycolytic flux, which varied depending on saccharide type and concentration. Overall, this study identified key mechanisms in carbohydrate metabolism - autoregulation and partial hierarchical control - and demonstrated that they are partly mediated by CcpA.

Towards virtual histology with X-ray grating interferometry.

Breast cancer is the most common type of cancer worldwide. Diagnosing breast cancer relies on clinical examination, imaging and biopsy. A core-needle biopsy enables a morphological and biochemical characterization of the cancer and is considered the gold standard for breast cancer diagnosis. A histopathological examination uses high-resolution microscopes with outstanding contrast in the 2D plane, but the spatial resolution in the third, Z-direction, is reduced. In the present paper, we propose two high-resolution table-top systems for phase-contrast X-ray tomography of soft-tissue samples. The first system implements a classical Talbot-Lau interferometer and allows to perform ex-vivo imaging of human breast samples with a voxel size of 5.57 µm. The second system with a comparable voxel size relies on a Sigray MAAST X-ray source with structured anode. For the first time, we demonstrate the applicability of the latter to perform X-ray imaging of human breast specimens with ductal carcinoma in-situ. We assessed image quality of both setups and compared it to histology. We showed that both setups made it possible to target internal features of breast specimens with better resolution and contrast than previously achieved, demonstrating that grating-based phase-contrast X-ray CT could be a complementary tool for clinical histopathology.

Co-utilization of saccharides in mixtures: Moving toward a new understanding of carbon metabolism in Streptococcus thermophilus.

The lactic acid bacterium Streptococcus thermophilus is widely used in food production, notably in yogurt fermentation. It evolved under highly specific ecological conditions, resulting in its ability to efficiently metabolize lactose, the main saccharide in milk. However, when used in sweetened dairy products or plant-based products, S. thermophilus may encounter other saccharides (i.e. alone or in mixtures). To date, the bacterium's metabolic capacities in such contexts have been poorly characterized. Here, we explored saccharide utilization by 39 S. thermophilus strains. Using in silico analysis, we discovered that the identity and structure of saccharide utilization genes are conserved across strains, and we identified six saccharides that might be metabolized. Although underlying genetic variability was low, strains nonetheless displayed differences in growth when supplied with different saccharides: lactose, sucrose, fructose, and glucose. Interestingly, we found that strains preferentially used lactose and sucrose in tandem when given saccharide mixtures. Furthermore, we uncovered some main potential drivers of saccharide metabolism in S. thermophilus. Notably, the sucrose transporter ScrA is also responsible for importing glucose. Overall, this research has yielded useful findings that can help the development of new fermented foods, including plant-based products, in which sucrose may serve as a major carbon source.

Assessing the use of nanoimmobilized laccases to remove micropollutants from wastewater.

Enzymes immobilization is a useful way to allow enzyme reuse and increase their stability. A high redox potential laccase from Trametes versicolor (TvL) and a low redox potential, but commercially available low-cost laccase from Myceliophthora thermophila (MtL), were successfully immobilized and co-immobilized onto fumed silica nanoparticles (fsNP). Enzyme loads of 1.78 ± 0.07, 0.69 ± 0.03, and 1.10 ± 0.01 U/mg fsNP were attained for the optimal doses of TvL, MtL, and co-immobilized laccases, respectively. In general, the laccase-fsNP conjugates showed a higher resistance against an acidic pH value (i.e., pH 3), and a higher storage stability than free enzymes. In addition, immobilized enzymes exhibited a superior long-term stability than free laccases when incubated in a secondary effluent from a municipal wastewater treatment plant (WWTP). For instance, the residual activity after 2 weeks for the co-immobilized laccases and the mixture of free laccases were 40.2 ± 2.5% and 16.8 ± 1.0%, respectively. The ability of the laccase-fsNP to remove a mixture of (14)C-bisphenol A (BPA) and (14)C-sodium diclofenac (DCF) from spiked secondary effluents was assessed in batch experiments. The catalytic efficiency was highly dependent on both the microbial source and state of the biocatalyst. The high redox potential TvL in free form attained a four-fold higher percentage of BPA transformation than the free MtL. Compared to free laccases, immobilized enzymes led to much slower rates of BPA transformation. For instance, after 24 h, the percentages of BPA transformation by 1000 U/L of a mixture of free laccases or co-immobilized enzymes were 67.8 ± 5.2 and 27.0 ± 3.9%, respectively. Nevertheless, the use of 8000 U/L of co-immobilized laccase led to a nearly complete removal of BPA, despite the unfavorable conditions for laccase catalysis (pH ~ 8.4). DCF transformation was not observed for any of the enzymatic systems, showing that this compound is highly recalcitrant toward laccase oxidation under realistic conditions.

[Paediatric emergency: creation of an independent nurse practitioner consultation]. / Pédiatrie. 4. Urgences pédiatriques: création d'une consultation infirmière indépendante.

Consultations in the Paediatric Emergency Department (PED) continue to climb regularly. Emergency Nurse Practitioner consultations have long been created in the English speaking countries. Since January 2013, an indepen- dent nurse consultation, under delegated medical responsibility, exists in the multidisciplinary PED of the Children's Hospital of Lausanne. The mean consultation time is the same as the medical consultation and the overall waiting time hasn't decreased yet. But a well definite working frame, a systematic approach, as well as the continual medical supervision possibility, make it a safe, efficient and appreciated consultation, by both patients and professionals.

Joint Alpharma-Beef Species Symposium: considerations on puberty in replacement beef heifers.

Many important changes occur throughout reproductive development in beef heifers, including during the prenatal, early postnatal, and peripubertal periods. Wave-like patterns of follicular development have been observed in heifer calves as early as 2 wk of age. Some dramatic changes occur from about 2 to 5 mo of age, most notably the transient increase in LH secretion. Most components of the hypothalamic-pituitary-ovarian axis are fully competent by approximately 5 to 6 mo of age. Peripubertal changes include increases in LH secretion, estradiol production, follicular development, and reproductive tract size. Eventually, the process reaches the point that the initial ovulation is achieved. Heifers that reach puberty and experience multiple estrous cycles before the onset of their initial breeding season have a greater probability for early conception and optimal lifetime productivity. Attainment of puberty typically occurs at around 12 to 14 mo of age in beef heifers but varies greatly. Genetic differences and environmental factors contribute to this variation. In typical U.S. cow-calf operations, calves are generally weaned at approximately 200 d of age. The impact of postweaning management on age at puberty in heifers has been demonstrated, and there is considerable flexibility in the timing of gain from weaning to breeding. However, even when heifers are grown to the desired BW before the start of breeding, there remains a pronounced variation in the timing of puberty, which impacts pregnancy rates. Less attention has been focused on the impact of preweaning management on age at puberty. Heifer calves with increased growth rates from birth to weaning have reached puberty at earlier ages. Precocious puberty has also been induced in a majority of heifers with early weaning and feeding a high-concentrate diet. Nutritional control during early maturation in heifers exerts a substantial influence on the timing of puberty. Understanding the mechanisms involved in reproductive development increases our ability to effectively manage replacement beef heifers for optimal reproductive performance.

[Disabled elderly people waiting for institutionalization to a hospital ward: prospective study in the administrative district of Strasbourg (France)]. / Personnes âgées dépendantes en attente d'entrée en EHPAD depuis un service hospitalier: étude prospective sur la communauté urbaine de Strasbourg, France.

OBJECTIVES: Our aim was to estimate the number of non-satisfied instutionalization requests for inpatients and to describe the strategies elaborated to compensate for the waiting time. METHODS: This prospective follow-up study concerning all requests for institution admission for inpatients aged 75 years or older hospitalized in acute care and rehabilitation wards. Descriptive data were gathered throughout the social support process conducted during the hospitalization. A three months follow-up was conducted. RESULTS: Among 5200 hospitalizations, a social support process was initiated for 270 patients aged 75 years and over. Two thirds of the sample were women (n=163). Mean age was 82 years. Fifty-two percent of the subjects met the criteria for iso-resource grades (IRG) 1 to 2 and 90% in IRG 1 to 4. The mean length of hospitalized stay (MLOS) was 56.8+/-10.2 days; the MLOS of unjustified stay of 23.5+/-5.6 (n=222). The average time before the social worker was informed of the patient's situation was 13.6+/-2.0 days; in addition, the time required to establish the administrative documents necessary for initiation of the social support progress was 15.0+/-1.8. The principal reasons for social support were physical dependence (77%), mental dependence (60%), insufficient family support (36%) and/or disease progression (21%). At three months, 104 patients were institutionalized, 128 were still on institution waiting list (in hospital: 48%; at home: 16%) and 38 had died (14%). The estimated annual institutional deficit for disabled elderly people was 512 beds. CONCLUSION: In light of demographical perspectives, an overall re-organization of the geriatric network is absolutely necessary. A simple increase in the capacity to fulfil the institutional beds deficit would be insufficient.

Decreasing the interval between GnRH and PGF2alpha from 7 to 5 days and lengthening proestrus increases timed-AI pregnancy rates in beef cows.

Four experiments were conducted in postpartum beef cows to evaluate the influence of reducing the interval from GnRH to PGF(2alpha) from 7 to 5d in a Select-Synch + CIDR or CO-Synch + CIDR estrous synchronization program. In Expt 1, cows (n=156) were treated with either a 7 or 5d Select-Synch + CIDR program. A second PGF(2alpha) treatment was given to all cows in all experiments at 12h after the initial PGF(2alpha) (to ensure that luteolysis occurred with the 5d program). Estrous response, interval to estrus, conception rate, and first service AI pregnancy rates were similar between treatments. In Expt 2, cows (n=223) were treated with either a 7 or 5d CO-Synch + CIDR program, with timed-AI concomitant with GnRH at 60 h after PGF(2alpha). Timed-AI pregnancy rates were similar between treatments. In Expt 3 (n=223) and 4 (n=400) cows were treated with either a 7 or 5d CO-Synch + CIDR program with timed-AI concurrent with GnRH at either 60 h (7d) or 72 h (5d) after CIDR withdrawal. Timed-AI pregnancy rates were 13.3% (P<0.05; Expt 3) and 9.1% (P<0.05; Expt 4) greater for the 5 than 7d program. In conclusion, timed-AI pregnancy rates were improved with a 5d CO-Synch + CIDR program with timed-AI at 72 h after CIDR withdrawal, compared to a 7d CO-Synch + CIDR program with timed-AI at 60 h after CIDR withdrawal.

Steroidogenic changes and steady state amount of messenger RNA encoding steroidogenic enzymes, gonadotropin receptors and cell-death signalling in the dominant ovarian follicle during estradiol-induced atresia in cattle.

Changes in steroidogenic function and associated gene expression were characterized in dominant ovarian follicles (DF) of cattle where follicles were induced to become atretic by systemic administration of estradiol benzoate (EB). In experiment 1, follicular fluid (FF) steroid concentrations in the DF were measured at 12-hourly time points for 48 h in heifers treated with 1 mg EB i.m./500 kg body weight (EB; n=20) as compared with untreated controls (C; n=19). Treatment with EB promoted a transient reduction in circulating FSH, a rapid (12 h) and sustained reduction in FF estradiol, a rapid (12 h) but transient reduction in FF progesterone and a delayed (36 h) increase in FF testosterone concentrations. In experiment 2, whole follicular wall tissue was collected from DF of mature non-lactating cows allocated to a 0 h control group (0 HC: n=7), a 24h control group (24 HC; n=7) or an EB-treated group where tissue was collected 24 h after administration of 1 mg EB i.m./500 kg body weight (EB; n=8). As for experiment 1, EB promoted a transient reduction in circulating FSH, a pronounced reduction in FF estradiol and a smaller but significant reduction in FF progesterone concentrations. Semi-quantitative RT-PCR on follicular wall tissue revealed that the loss in estrogen activity at 24 h after EB was associated with two-fold reduction in aromatase mRNA, with an apparent acceleration in loss of 17alpha-hydroxylase mRNA. Expression of genes for gonadotropin receptors (LHR and FSHR) and a cell-death signalling pathway (Fas antigen and Fas ligand) were unchanged during the initial 24h of EB-induced atresia. These results suggest that EB initiates atresia in dominant ovarian follicles through a rapid suppression of follicular estradiol synthesis, an effect associated with down-regulation of the aromatase gene. A transient suppression in circulating FSH following administration of EB appears to have initiated these events, and it is suggested that subsequent processes involved in atresia follow this loss in estrogenic function.

Influence of premature induction of a luteinizing hormone surge with gonadotropin-releasing hormone on ovulation, luteal function, and fertility in cattle.

We tested the hypothesis that luteal function and fertility would be reduced in cattle induced to ovulate prematurely compared with those ovulating spontaneously. Estrus was synchronized in 56 beef cows (24 that were nonlactating and 32 that were nursing calves). At 6.4 +/- 0.1 d after estrus, all follicles > or = 5 mm were aspirated (day of aspiration = d 0) with a 17-gauge needle using the ultrasound-guided transvaginal approach. On d 1.5 and 2, cows were administered 2 luteolytic doses of PGF2alpha. Ovarian structures were monitored by transrectal ultrasonography from d -2 to 12, or ovulation. Emergence of a new follicular wave occurred on d 1.7 +/- 0.1. When the largest follicle of the newly emerged wave was 10 mm in diameter (d 4.8 +/- 0.1), cows were assigned on an alternating basis to receive 100 microg of GnRH (GnRH-10; n = 29) to induce ovulation or, upon detection of spontaneous estrus, to the spontaneous (SPON) treatment (n = 24). Cows were bred by AI at 12 h after GnRH (GnRH-10) or 12 h after the onset of estrus (SPON) as detected using an electronic surveillance system. Blood samples were collected every other day beginning 2 d after ovulation until pregnancy diagnosis 30 d after AI. Ovulation and AI occurred in 29/29 cows in the GnRH-10 and in 24/24 cows in the SPON treatment. Ovulation occurred later (P < 0.05) in the SPON (d 7.7 +/- 0.1) than GnRH-10 (d 6.8 +/- 0.1) treatment. Double ovulations were detected in 47% of cows, resulting in 1.5 +/- 0.1 ovulations per cow. Diameters of the ovulatory and the second ovulatory (in cows with 2 ovulations) follicles were greater (P < 0.05) in the SPON (12.0 +/- 0.3 mm and 10.5 +/- 0.4 mm, respectively) than in the GnRH-10 (10.7 +/- 0.1 mm and 9.2 +/- 0.3 mm) treatment. Cross-sectional areas of luteal tissue and plasma concentrations of progesterone during the midluteal phase were greater (P < 0.05) in the SPON (3.62 +/- 0.2 cm2 and 6.4 +/- 0.3 ng/mL) than in the GnRH-10 (3.0 +/- 0.2 cm2 and 5.4 +/- 0.2 ng/mL) treatment. The conception rate to AI in the SPON (100%) treatment was greater (P < 0.05) than in the GnRH-10 (76%) treatment. The animal model used in this study resulted in unusually high conception rates and double ovulations. In conclusion, premature induction of the LH surge reduced the diameter of ovulatory follicle(s), the luteal function, and the conception rate to AI.

Effect of timing of feeding a high-concentrate diet on growth and attainment of puberty in early-weaned heifers.

Precocious puberty (<300 d of age) can be successfully induced in a majority of heifers with early weaning and continuous feeding of a high-concentrate diet. The objective of this experiment was to determine the relative effects of timing of feeding a high-concentrate diet on age at puberty in early-weaned heifers. Sixty crossbred Angus and Simmental heifer calves were weaned at 112 +/- 2 d of age and 155 +/- 3 kg of BW and were fed a receiving diet for 2 wk. Heifers were blocked by age and BW, and assigned randomly to receive a high-concentrate (60% corn; H) or control (30% corn; C) diet during phase 1 (mean age 126 to 196 d) and H or C during phase 2 (mean age 196 to 402 d), resulting in 4 treatments (HH, n = 15; HC, n = 15; CH, n = 15; and CC, n = 15). Blood samples were collected weekly beginning at a mean age of 175 d and assayed for progesterone concentration to determine age at puberty. After 56 d on the experimental diets, BW of heifers fed the H diet during phase 1 were greater (P < 0.05) than those of heifers fed the C diet (mean age of 182 d; treatment x mean age, P < 0.01). After 70 d on the new diets (mean age of 266 d), heifers fed the H diet during phase 2 reached heavier BW (P < 0.05) than heifers fed the C diet, when compared within phase 1 diet groups (HH > HC; CH > CC). Body weights in HC and CH treatments differed from a mean age of 169 through 238 d, after which BW did not differ between these treatments. The ADG over the entire experimental period was greatest for the HH treatment (1.2 +/- 0.04 kg/d; P < 0.05), followed by the HC and CH treatments (1.0 +/- 0.03 and 1.0 +/- 0.02 kg/d, respectively), which were not different, and the CC treatment gained the least (0.7 +/- 0.04 kg/d; P < 0.05). Precocious puberty occurred in 67, 47, 47, and 20% of heifers in the HH, HC, CH, and CC treatments, respectively (HH > CC; P < 0.05). Mean age at puberty for the HH and HC treatments (271 +/- 17 and 283 +/- 17 d of age, respectively) was earlier (P < 0.05) than for the CC treatment (331 +/- 11 d of age). Age at puberty in the CH treatment (304 +/- 13 d of age) was intermediate to and not different from the other treatments. Heifers fed the H diet during phase 1 attained puberty earlier (P < 0.05) than heifers fed the C diet during phase 1. In conclusion, increasing dietary energy intake in early-weaned heifers, through feeding a high-concentrate diet from 126 to 196 d of age, decreased age at puberty regardless of the diet fed after 196 d of age.

Induction of precocious puberty in heifers I: enhanced secretion of luteinizing hormone.

In beef heifers weaned between 3 and 4 mo of age and fed a high-concentrate diet, approximately 50% reach puberty before 300 d of age (precocious puberty). The objectives of this experiment were 1) to determine whether precocious puberty could be induced experimentally by weaning heifers early and feeding a high-concentrate diet, and 2) to determine the dynamics of secretion of LH associated with precocious puberty. Crossbred Angus and Simmental heifer calves were weaned at 73 +/- 3 d of age and 115 +/- 3 kg of BW and fed a high-concentrate (60% corn; HI, n = 9) or control diet (30% corn; CONT, n = 9). Heifers were fed individually, and target BW gains were 1.50 and 0.75 kg/d for the HI and CONT treatments, respectively. Heifers were weighed every 2 wk. Blood samples were collected weekly and assayed for progesterone concentration to determine age at puberty. Serial blood samples were collected at 20-min intervals for 24 h at mean ages of 102, 130, 158, 172, 190, 203, 217, 231, and 259 d and assayed for LH concentration to evaluate the dynamics of secretion of LH. Heifers fed the HI diet exhibited greater BW gain (P < 0.01) than CONT heifers (1.27 +/- 0.05 vs. 0.85 +/- 0.05 kg/d, respectively). As a result, BW in the HI treatment was greater (P < 0.01) than in the CONT treatment by 188 d of age and remained different through the end of the experiment. Precocious puberty occurred in 8 of 9 heifers fed the HI diet and 0 of 9 heifers fed the CONT diet. Age at puberty was reduced in the HI (P < 0.01) compared with the CONT heifers (262 +/- 10 vs. 368 +/- 10 d of age, respectively). Body weight at puberty was also reduced in the HI (P < 0.05) compared with the CONT treatment (327 +/- 17 vs. 403 +/- 23 kg, respectively). Heifers attaining puberty during the experiment continued with subsequent luteal phases as evidenced by cyclic patterns of progesterone concentrations. Frequency of pulses of LH (pulses/24 h) increased with age (P < 0.01) for both treatments. Heifers in the HI treatment exhibited a greater number of pulses of LH (P < 0.01) than those in the CONT treatment by 190 d of age and in all subsequent collection periods (treatment x age, P < 0.05). Mean LH concentrations also increased with age (P < 0.01) for both treatments but did not differ between treatments. In conclusion, precocious puberty induced by early weaning and feeding of a high-concentrate diet is preceded by increasing frequency of pulses of LH.

Induction of precocious puberty in heifers III: hastened reduction of estradiol negative feedback on secretion of luteinizing hormone.

Precocious puberty (<300 d of age) can be induced in beef heifers by early weaning and feeding a high-concentrate diet. The objective of this experiment was to determine whether precocious puberty occurs as a result of a hastened reduction of estradiol negative feedback on secretion of LH. Thirty crossbred Angus and Simmental heifers were weaned at 83 +/- 2 d of age and 114 +/- 3 kg of BW, blocked by BW, and randomly assigned to receive a high-concentrate (60% corn; H) or control (30% corn; C) diet and to receive ovariectomy (OVX), OVX plus an estradiol implant (OVXE), or to remain intact (INT). Residual ovarian tissue after OVX necessitated withdrawal of 6 heifers during the course of the experiment, resulting in the following treatment groups: OVX-C, n = 3; OVX-H, n = 5; OVXE-C, n = 4; OVXE-H, n = 2; INT-C, n = 5; INT-H, n = 5. To determine concentrations of progesterone and estradiol, blood samples were collected weekly beginning at a mean age of 160 d. To characterize LH concentrations, serial blood samples were collected at 12-min intervals for 12 h at mean ages of 119, 149, 188, 217, 246, 281, 323, 365, 407, and 449 d. By a mean age of 202 d, heifers fed the H diet were heavier (P < 0.05) than those fed the C diet. Heifers in the INT-H treatment attained puberty earlier (P < 0.05) than in the INT-C treatment (275 +/- 30 vs. 385 +/- 14 d of age, respectively). Overall mean concentrations of estradiol did not differ between OVXE-H and OVXE-C, between INT-H and INT-C, or between OVXE and INT treatments. The OVX treatments exhibited greater LH pulse frequency than the OVXE and INT treatments by the first serial blood collection (treatment x age, P < 0.05). The frequency of LH pulses was greater (P < 0.05) in the INT-H than the INT-C treatment by a mean age of 246 d and was greater (P < 0.05) in the OVXE-H than the OVXE-C treatment by a mean age of 281 d. In the OVXE-H treatment, LH secretion increased and subsequently "escaped" from estradiol negative feedback (detection of > or = 1 LH pulse/h) earlier (P < 0.05) than in the OVXE-C treatment (307 +/- 30 and 420 +/- 21 d of age, respectively). It is concluded that advancing the reduction of estradiol negative feedback on secretion of LH is the mechanism by which early weaning and feeding a high-concentrate diet results in precocious puberty in heifers.

Induction of precocious puberty in heifers II: advanced ovarian follicular development.

Precocious puberty can be induced in a majority of heifers weaned early and fed a high-concentrate diet. The objective of this experiment was to determine whether induction of precocious puberty is associated with an acceleration of ovarian maturation in heifers. Crossbred Angus and Simmental heifer calves were weaned at 104 +/- 2 (n = 18; early weaned) or 208 +/- 3 (n = 10; normal-weaned, NW) d of age. The early weaned heifers were fed a high-concentrate (60% corn; EWH, n = 9) or control diet (30% corn; EWC, n = 9). The NW heifers were also fed the control diet after weaning. Daily transrectal ultrasonography was performed to characterize a complete follicular wave beginning at a mean age of 126, 161, 196, 224, and 252 (EWH and EWC), or 224 and 252 (NW) d. Blood samples were collected daily during periods of ultrasonography to determine estradiol concentrations and weekly beginning at mean ages of 153 (EWH and EWC) or 216 (NW) d to be analyzed for progesterone concentrations. Heifers in the EWH treatment were heavier (P < 0.01) than EWC heifers from a mean age of 175 d through the end of the study (treatment x age; P < 0.05). Body weights did not differ between EWC and NW. At mean ages of 196 and 224 d, the maximum diameter of the dominant follicle (MaxDF) was greater (P < 0.05) in EWH than EWC heifers. At a mean age of 224 d, MaxDF was greater (P < 0.05) in EWC than NW heifers but was not different by a mean age of 252 d. All EWH, 5 of 9 EWC, and 5 of 10 NW heifers attained puberty at less than 300 d of age (precocious puberty). Age at puberty was less (P < 0.05) in EWH (252 +/- 9 d) than in EWC and NW (308 +/- 26 and 330 +/- 25 d, respectively) treatments. Across all heifers, MaxDF and duration of follicular waves increased with age (P < 0.05), mean number of follicles during follicular waves decreased with age (P < 0.05), and peak concentrations of estradiol during follicular waves increased until a mean age of 224 d. To further characterize aspects of precocious puberty, heifers were compared across treatments between those that experienced precocious puberty and those that did not. In heifers that experienced precocious puberty, BW at puberty was less (P < 0.01) and MaxDF, follicular wave duration, and peak estradiol concentrations were greater (P < 0.05) compared with heifers that did not experience precocious puberty. Ovarian maturation was accelerated in heifers that were weaned early and fed a high-concentrate diet and was associated with precocious onset of puberty.

Estradiol benzoate delays new follicular wave emergence in a dose-dependent manner after ablation of the dominant ovarian follicle in cattle.

Administration of estradiol benzoate (EB) induces atresia of the dominant follicle (DF) in the ovaries of cattle within 36 h but emergence of a new wave of follicular development is delayed by 3-5 days. The present study investigated the role of EB in determining timing of emergence of a new follicular wave after removing the influence of the DF. At 6.4+/-0.2 days after ovulation in Angus and Angus/Simmental cattle (n=26), aged 4.9+/-0.6 years and weighing 634+/-20 kg, all ovarian follicles > or =5mm in diameter were aspirated with a 17-gauge needle using an ultrasound-guided transvaginal approach (Day 0 or Hour 0) and animals immediately received 0 (0EB), 1 (1EB), 2 (2EB) or 4 (4EB) mg EB i.m./500 kg body weight (n=6 or 7 per treatment). Ovarian structures were monitored by ultrasonography on a daily basis until emergence of a new wave of follicular development. Concentrations of estradiol (E2) were different among all treatments between Hours 24 and 72, increasing (P<0.01) with greater doses of EB administered. Hour of peak follicle-stimulating hormone (FSH) was 29.3+/-4.0, 53.3+/-4.5, 81.1+/-15.5, and 91.4+/-8.2 for the 0EB, 1EB, 2EB, and 4EB treatments, respectively, and emergence of a new wave of follicular development occurred on Days 1.5+/-0.2, 3.3+/-0.3, 4.0+/-0.6 and 4.4+/-0.4, respectively. Timing of peak FSH and emergence of a new wave of follicular development was earliest (P<0.05) in the 0EB treatment, similar (P>0.1) among the 1EB and 2EB treatments, and most delayed (P<0.05) in the 4EB treatment when compared to the 0EB or 1EB treatments. The overall mean interval from peak FSH to emergence of a new wave of follicular development was 15.7+/-3.3 h and was not affected by treatment. Concentrations of E2 at 24 h before new emergence were not different among EB-treated animals (20.2+/-5.5 pg/ml), but lower (P<0.01) in the 0EB treatment (1.6+/-0.2 pg/ml). In a dose-dependent manner, EB delayed the pre-emergence surge in FSH that stimulates new follicular development after the DF has ceased to be functional. The importance of using an 'optimal' dose of EB in hormonal regimens using this agent to strategically regulate follicular development is emphasized by the outcomes of this study.

The Arabidopsis HUELLENLOS gene, which is essential for normal ovule development, encodes a mitochondrial ribosomal protein.

The HUELLENLOS (HLL) gene participates in patterning and growth of the Arabidopsis ovule. We have isolated the HLL gene and shown that it encodes a protein homologous to the L14 proteins of eubacterial ribosomes. The Arabidopsis genome also includes a highly similar gene, HUELLENLOS PARALOG (HLP), and genes for both cytosolic (L23) and chloroplast ribosome L14 proteins. Phylogenetic analysis shows that HLL and HLP differ significantly from these other two classes of such proteins. HLL and HLP fusions to green fluorescent protein were localized to mitochondria. Ectopic expression of HLP complemented the hll mutant, indicating that HLP and HLL share redundant functions. We conclude that HLL and HLP encode L14 subunits of mitochondrial ribosomes. HLL mRNA was at significantly higher levels than HLP mRNA in pistils, with the opposite pattern in leaves. This differential expression can explain the confinement of effects of hll mutations to gynoecia and ovules. Our elucidation of the nature of HLL shows that metabolic defects can have specific effects on developmental patterning.

Recent progress in reconstructing angiosperm phylogeny.

In the past year, the study of angiosperm phylogeny has moved from tentative inferences based on relatively small data matrices into an era of sophisticated, multigene analyses and significantly greater confidence. Recent studies provide both strong statistical support and mutual corroboration for crucial aspects of angiosperm phylogeny. These include identifying the earliest extant lineages of angiosperms, confirming Amborella as the sister of all other angiosperms, confirming some previously proposed lineages and redefining other groups consistent with their phylogeny. This phylogenetic framework enables the exploration of both genotypic and phenotypic diversification among angiosperms.

SHORT INTEGUMENTS 2 promotes growth during Arabidopsis reproductive development.

The short integuments 2 (sin2) mutation arrests cell division during integument development of the Arabidopsis ovule and also has subtle pleiotropic effects on both sepal and pistil morphology. Genetic interactions between sin2 and other ovule mutations show that cell division, directionality of growth, and cell expansion represent at least partially independent processes during integument development. Double-mutant analyses also reveal that SIN2 shares functional redundancy with HUELLENLOS in ovule primordium outgrowth and proximal-distal patterning and with TSO1 in promotion of normal morphological development of the four whorls of primary floral organs. All of these observations are consistent with SIN2 being a promoter of growth and cell division during reproductive development, with a primary role in these processes during integument development. On the basis of the floral pleiotropic effects observed in a majority of ovule mutants, including sin2, we postulate a relationship between ovule genes and the evolutionary origin of some processes regulating flower morphology.

TSO1 is a novel protein that modulates cytokinesis and cell expansion in Arabidopsis.

Previous analyses of tso1 mutants revealed a loss of control of directional cellular expansion and coordination of growth of adjacent cells, and defects in karyokinesis and cytokinesis. We isolated TSO1 using a map-based approach, and show that it is a member of a family of at least three genes in Arabidopsis. Consistent with the mutant phenotype, TSO1 transcript was most abundant in flowers, where it accumulated to the highest levels in developing ovules and microspores. The putative TSO1 protein has two cysteine-rich regions that are similar to the CXC domains of a variety of proteins from plants and animals, including a class of kinesins involved in chromosome segregation, and enhancer of zeste-type proteins. Visualization of TSO1-fusion proteins indicated that TSO1 is a nuclear protein. The tso1 mutant phenotypes and the novelty of the TSO1 sequence suggest the existence of previously unknown participants in regulation of directional processes in eukaryotic cells.

INNER NO OUTER regulates abaxial- adaxial patterning in Arabidopsis ovules.

The Arabidopsis INNER NO OUTER (INO) gene is essential for formation and asymmetric growth of the ovule outer integument. INO encodes a member of the newly described YABBY family of putative transcription factors that contain apparent Cys(2)-Cys(2) zinc-finger domains and regions of similarity to the high mobility group (HMG) transcription factors. In wild-type plants, INO is expressed specifically on one side of the central region of each ovule primordium in the cells that give rise to the outer integument. Alterations in the INO expression pattern in mutant backgrounds implicate INO as a positive regulator of its own expression, and ANT, HLL, BEL1, and SUP as direct or indirect negative regulators that help to establish the spatial pattern of INO expression. We hypothesize that INO is necessary for polarity determination in the central part of the ovule. Maintenance of polarity in other parts of ino ovules indicates the existence of additional regulators and provides further evidence that the ovule is a compound structure.