Zygote donor nitrogen metabolism and in vitro embryo culture perturbs in utero development and IGF2R expression in ovine fetal tissues

Tests were made of the effects of altering nitrogen metabolism in zygote donor ewes on fetal development and expression of the gene encoding the type II insulin-like growth factor receptor (IGF2R) following the transfer of ovine embryos cultured from these zygotes, either in the absence or presence of serum. Zygotes, recovered from superovulated ewes (32 on a urea supplemented (30 g urea/kg) diet (high N) and 32 on a control diet (low N)) 36 h after intrauterine AI using semen from a single sire, were cultured for 5 days in synthetic oviductal fluid (SOF) media either with BSA and amino acids (SOF-) or with 10% (v/v) steer serum (SOF+). In total, 166 embryos, including 30 in vivo controls, were transferred singly at day 6 post-AI to synchronous recipients and the products of conception recovered at day 125 of gestation. Elevated plasma urea concentrations in zygote donors were associated with accelerated early embryo development, low pregnancy rates (16%) for embryos from the high N, SOF+ treatment, and significantly influenced fetal development and the expression of IGF2R in the fetal heart at day 125 of gestation. Importantly, the culture of sheep zygotes under serum-free conditions led to a high incidence of aberrant conceptus development and IGF2R expression. Consequently, maternal nitrogen metabolism prior to zygote recovery and in vitro culture can influence fetal development and the expression of an imprinted gene following embryo transfer, and these data support the notion that environmental effects on the follicle-enclosed oocyte may contribute to the etiology of the Large Offspring Syndrome.     

The mechanism of trichome resistance in Anaphalis margaritacea to the meadow spittlebug Philaenus spumarius

This study demonstrated how simple, lanate trichomes on the stem of Anaphalis margaritacea (D.C.) inhibit feeding by young nymphs of Philaenus spumarius (L.). Nymphs were caged on stems which were subsequently sectioned and examined for feeding marks. The lack of stylet punctures at sites where nymphs were unable to feed indicated that trichomes prevented the start of stylet penetration rather than interfering with the progress of the stylets after penetration had begun. Trichome resistance declined progressively with spittlebug instar from complete resistance to first instar nymphs through complete absence of resistance to fifth instar nymphs. Discriminant analysis of the effect of number of trichomes per mm², vertical density, and height on trichome resistance to third instar nymphs revealed that trichome height was the critical factor in resistance. Variation in the percent of nymphs in each instar able to feed when caged on the stem was (1) positively correlated with degree to which the average rostrum length of the instar exceeded the trichome height, (2) independent of trichome density, which was uniformly high on all stems. Trichomes prevented stylet penetration where trichome height exceeded the length of the nymph's rostrum, probably because the necessary stimuli were not received when the sensilla on the tip of the rostrum could not touch the surface of the stem. A similar mechanism may explain resistance of simple trichomes to crop pests in the Cicadellidae.

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Résumé Ce travail montre comment des trichomes simples et laineux sur les tiges d'Anaphalis margaritacea (D.C.) inhibent la prise d'aliments par les jeunes larves de Philaenus spumarius L. Les larves ont été encagées sur des tiges qui ont été ultérieurement coupées et les piqûres alimentaires dénombrées. L'absence de piqûres par les stylets aux lieux où les larves ont été incapables de s'alimenter a montré que les trichomes ont empêché le début de la pénétration des stylets plus qu'ils n'ont entravé leur progression après le début de la pénétration. La résistance des trichomes diminue progressivement: totale pour le premier stade larvaire, elle est devenue nulle au cinquième stade. Une analyse discriminante des effets de la densité des trichome par mm², de la densité verticale, de leur hauteur sur la résistance des trichomes aux larves de 3ème stade, a révélé que la hauteur du trichome est le facteur de résistance. La variation dans le pourcentage de larves de chaque stade encagées sur la tige, capables de consommer, présentait une relation positive avec le degré par lequel la longueur du rostre dépasse la hauteur du trichome, quelle que soit la densité des trichomes qui est toujours élevée sur toutes les tiges. Les trichomes ont empêché la pénétration des stylets quand leur hauteur a dépassé la longueur du rostre, probablement parce que les stimuli nécessaires n'ont pas été reçus quant l'extrémité du rostre ne pouvait pas atteindre la surface de la tige. Un mécanisme semblable peut expliquer la résistance de trichomes simples aux Cicadellidae.

     

pULB113, an RP4::mini-Mu plasmid, mediates chromosomal mobilization and R-prime formation in Erwinia amylovora, Erwinia chrysanthemi, and subspecies of Erwinia carotovora

The RP4::mini-Mu plasmid pULB113, transferred from Escherichia coli strain MXR, was stable and transfer proficient in Erwinia amylovora strain EA303, E. carotovora subsp. atroseptica strain ECA12, E. carotovora subsp. carotovora strain ECC193, and E. chrysanthemi strain EC183. The plasmid mobilized an array of Erwinia sp. chromosomal markers (E. amylovora: his+,ilv+,rbs+,ser+,thr+;E. chrysanthemi:arg+,his+,ilv+,leu+; E. carotovora subsp. atroseptica: arg+,gua+,leu+,lys+,pur+,trp+; E. carotovora subsp. carotovora: arg+,gua+,leu+,lys+,out+[export of enzymes],pur+,trp+), suggesting random interactions of the plasmid with the chromosomes. In E. carotovora subsp. carotovora, pULB113-mediated two-factor crosses revealed linkage between three auxotrophic markers and the out loci. The export of pectate lyase, polygalacturonase, and cellulase and the maceration of potato tuber tissue occurred with Out+, but not Out-, strains of E. carotovora subsp. carotovora, indicating the importance of enzyme export in plant tissue maceration. Erwinia sp. donors harboring pULB113 complemented mutations in various biosynthetic and catabolic genes (arg, gal, his, leu, met, pro, pur, thy) in Escherichia coli recA strains. Escherichia coli transconjugants harbored pULB113 primes as indicated by the cotransfer of Erwinia genes and pULB113 markers and a change in plasmid mass. Moreover, the PstI and SmaI cleavage patterns of selected pULB113 primes were different from those of pULB113. pULB113 primes carried DNA insertions ranging from 3 to about 160 kilobases. These findings indicate that pULB113 is useful for in vivo gene cloning and genetic analysis of various enterobacterial phytopathogens.     

Bacterial contamination of the small intestine is an important cause of occult malabsorption in the elderly.

A study was conducted to examine the contribution that occult malabsorption makes to malnutrition among elderly patients admitted to an acute geriatric ward. Malnutrition was defined by anthropometric, haematological, and biochemical measurements. Out of 490 patients, 55--many of whom had presented with non-gastrointestinal symptoms--were found to be malnourished. In 31 poor diet alone was probably the cause, but in the remaining 24 patients previously unrecognised malabsorption was detected. Of these patients, 17 were found to have bacterial contamination of the small bowel (nine with duodenojejunal diverticula, four after gastric surgery, and four diagnosed as contamination with "no sump"). In 10 patients contamination was confirmed by culture of intestinal juice: Escherichia coli was predominant in nine cases, and anaerobic organisms in one. Small bowel bacterial overgrowth without a "blind loop" or obvious underlying cause has not previously been fully proved in the elderly. Coeliac disease was detected in two patients, and chronic pancreatitis in two. Occult malabsorption is an important cause of malnutrition in the elderly. Such malnutrition not infrequently occurs in geriatric patients presenting to hospital with non-specific symptoms of physical deterioration.     

Superovulatory responses to eCG in llamas (Lama glama )

Llamas are copulation-induced single-ovulators, and multiple ovulation and embryo transfer (MOET) methods have not yet been developed for this species. Superovulatory responses to eCG given during an induced (Group A) or simulated (Group B) luteal phase were investigated using ultrasound to observe ovarian follicles and corpora lutea (CLs) and plasma progesterone was used to assess luteal function. Embryos were recovered nonsurgically. Group A (n = 19): donors were given 8 microg, im GnRH analogue (Day 0) to induce ovulation of a mature follicle, 1000 IU, im eCG (Day 7), and 250 microg PGF(2alpha) analogue (Day 9). Group B (n = 17): donors were given a subcutaneous progestagen implant (3 mg Norgestomet) at Days 0 to 7) and 1000 IU, im eCG (Day 5). When most (>65%) of the follicles in both Groups A and B had matured at 5 to 11 d post eCG, the donors were given 8 microg, im GnRH and mated once (n = 26) or twice within a 24-h interval (n = 10); embryos were recovered 6 to 9 d post ovulation. More follicles and corpora lutea were induced in Group B than in Group A, but a similar mean number of embryos were recovered (1.3 vs 1.6), and a similar proportion of donors yielded multiple embryos (35 vs 32%). The embryo recovery rate was similar for Groups A and B (39 and 37%), but it was higher (P < 0.001) with 2 (72%) rather than 1 (22%) mating, and it was negatively correlated with CL number (P < 0.05). Overall, 80% of the llamas had a precocious CL and elevated plasma progesterone concentrations when multiple follicles reached maturity. This was associated with increased subsequent superovulation and embryo recovery (P < 0.01). Peak plasma progesterone was positively correlated with the CL number (P < 0.05). From these results we conclude that superovulation may be achieved with eCG given during either an induced or a simulated luteal phase, that embryo recovery is improved following 2 matings rather than 1, and that MOET may indeed be feasible for use in the llama.