Failure of spermatozoa from T/t mice to fertilize in vitro is overcome by zona drilling

Failure of epididymal spermatozoa from T/t mutant mice, but not from t/t individuals, to fertilize oocytes in vitro was partially overcome by opening a small aperture in the zona pellucida with acidified Tyrode's solution to permit direct access of the spermatozoon to the vitellus. This study provides a model system to evaluate requirements for successful zona drilling in the treatment of human infertility and further insights into the effects of the t complex on sperm fertility.     

Use of diethyl squarate for the coupling of oligosaccharide amines to carrier proteins and characterization of the resulting neoglycoproteins by MALDI-TOF mass spectrometry

The 8-methoxycarbonyloctyl glycosides of GlcNAc, Gal1-4Glc, Fuc1-2Fuc1-3GalNac and Fuc1-2Gal1-3[Fuc1-4]GlcNac were converted to primary amines by reaction with neat ethylenediamine and then coupled to bovine serum albumin (BSA) using diethyl squarate as the connector. The average degree of incorporation of the sugar onto the protein, as well as the molecular weight distribution, could be conveniently determined using matrix assisted laser desorption ionization/time of flight (MALDI-TOF) mass spectrometry thus avoiding cumbersome structure-dependent colour-tests or analysis of cleaved ligand. The present coupling method has the advantages of proceeding under very mild conditions, yielding controlled incorporation values and can reliably be used for the coupling of very small amounts (mg) of oligosaccharide.This paper is dedicated to Sen-itiroh Hakomori on the occasion of his 65th birthday     

Molecular selection in apple for resistance to scab caused by Venturia inaequalis

Large-scale marker-assisted selection requires highly reproducible, consistent and simple markers. The use of genetic markers is important in woody plant breeding in general, and in apple in particular, because of the high level of heterozygosity present in Malus species. We present here the transformation of two RAPD markers, which we found previously to be linked to the major scab resistance gene Vf, into more reliable and reproducible markers that can be applied directly to apple breeding. We give an example of how the use of such markers can speed up selection for the introduction of scab resistance genes into the same plant, reducing labour and avoiding time-consuming test crosses. We discuss the nature and relationship of the scab resistance gene Vf to the one present in Nova Easygro, thought to be Vr.     

Ultrafast protein determinations using microwave enhancement

We present here microwave-based modifications of standard protein assays that dramatically reduce the time required to determine protein concentrations. Typical protein determinations involve incubation times ranging from 15–60 min. Microwave irradiation of specimens reduces this time requirement to 10–20 s without compromising accuracy or reliability. The remarkable speed with which protein determinations may be carried out using microwave enhancement greatly simplifies general laboratory procedures that depend on the estimation of protein concentrations. An erratum to this article is available at .     

Presence of double‐stranded RNAs and virus‐like particles in the entomopathogenic fungus Metarhizium anisopliae

Nucleic acids from 41 strains of Metarhizium anisopliae, obtained from different parts of the world were extracted and examined by electrophoresis. Strong bands of double‐stranded RNA (dsRNA) were detected in two isolates from Brazil, V215 and V291, which had, respectively, 13 and 9 distinct bands ranging in size from ca. 0.75 to 3.5 kb. Icosahedral virus‐like particles (VLPs) (ca. 33 nm in diameter) were observed by transmission electron microscopy in extracts of these isolates. The VLPs and dsRNA were both absent from a clone of the isolate V291 which had been subcultured successively on solid medium. Bioassays against the aphid Myzus persicae showed no detectable difference in virulence between the clone of V291 which contained dsRNA and the clone that did not.     

Essential trace elements in humans

Serum arsenic concentrations of persons suffering from renal failure and undergoing hemodialysis treatment (n=85) and of healthy controls (n=25) were determined by hydride-generation AAS technique after microwave digestion. The results were evaluated by comparing the values of both groups, considering physiological factors and individual data, as well as comorbid conditions of the hemodialysis (HD) patients. Serum arsenic levels were diminished in the patient group compared with controls (mean values 8.5±1.8 ng/mL vs 10.6±1.3 ng/mL). Furthermore, additional diseases within the hemodialysis group, particularly injuries of the central nervous system (CNS), vascular diseases, and cancer, were correlated to occasionally markedly decreased serum arsenic concentrations. It was concluded that arsenic homeostasis is disturbed by HD treatment and certain additional diseases. Desirable arsenic concentrations in the body seem to be reasonable. This consideration results in the conclusion that arsenic could play an essential role in human health. Thus, reference arsenic concentrations in different human tissues and body fluids should be established in order to recognize not only arsenic intoxication, but also arsenic deficiency. Perhaps arsenic deficiency contributes to the increased death risk of HD patients, and therefore, arsenic supplementations for patients with extremely low serum arsenic concentrations should be taken into account.     

Antibiotic production by strains of Gliocladium virens and its relation to the biocontrol of cotton seedling diseases

Strains of the fungal antagonist Gliocladium virens were separated into two distinct groups on the basis of secondary metabolite production in vitro. Strains of the ‘P’ group produced the antibiotics gliovirin and heptelidic acid but not the antibiotic gliotoxin and its companion, dimethylgliotoxin. Strains of the ‘Q’ group produced gliotoxin and dimethylgliotoxin but not gliovirin or heptelidic acid. Strains from both groups produced the antibiotic viridin and phytotoxin viridiol. Gliovirin was very inhibitory to Pythium ultimum but had no activity against Rhizoctonia solani, and strains that produce it were more effective seed treatment biocontrol agents of disease incited by P. ultimum. Conversely, gliotoxin was more active against R. solani than against P. ultimum, and strains that produced it were more effective seed treatments for controlling disease incited by R. solani. These results indicate that the antibiotic profiles of strains should be considered when screening strains for biocontrol efficacy, and that it may be necessary to treat seeds with a combination of strains in order to broaden the disease control spectrum.     

Reproductive status of cheetahs (Acinonyx jubatus) in North American Zoos: The benefits of physiological surveys for strategic planning

Under the mandate of a Species Survival Plan (SSP), reproductive status was assessed in 128 cheetahs maintained in 18 different institutions in North America. A mobile laboratory research team evaluated cheetahs using anesthesia, serial blood sampling, electroejaculation (males), and laparoscopy (females). Biomaterials were also collected for parallel studies of genetics, nutrition, and health. There was no mortality, and cheetahs were capable of reproducing naturally after these intense manipulatory examinations. No marked differences were observed in reproductive or endocrine characteristics between proven and unproven breeders. However, males consistently produced teratospermic ejaculates, and cheetah sperm were compromised in conspecific or heterologous in vitro fertilization systems. Structurally abnormal sperm were found to be filtered by the oocyte's zona pellucida. More than 80% of the females were anatomically sound, but morphological and endocrine evidence suggested that ～50% or more of the population may have had inactive ovaries at the time of the examination. Males ranging in age from 15 to 182 months produced spermic ejaculates, but motile sperm numbers/ejaculate and circulating testosterone concentrations were highest in males 60 to 120 months old. Parovarian cysts were observed in 51.5% of female cheetahs, but comparisons between proven and unproven subpopulations revealed that this abnormality likely had no influence on fertility. Fresh luteal tissue was not observed in any nonpregnant or nonlactating female, strongly suggesting that the cheetah is an induced ovulator. Overall survey results were discussed in the context of the etiology of reproductive inefficiency, especially with respect to the potential importance of biological versus management factors. Four high priority research areas in cheetah reproductive biology were identified: 1) continuous monitoring of ejaculate quality in the extant population, while studying the impact of pleiomorphisms on fertility; 2) determining the potential relationship between libido and androgen production (excretion) in males; 3) confirming the extent of cyclic, or acyclic, ovarian activity in females; and 4) continued development of assisted reproductive techniques for enhancing man-agement. In summary, a multidisciplinary, multi-institutional survey coordinated through the SSP is both possible and useful for generating a physiological and health database beneficial to driving further research and management initiatives. © 1993 Wiley-Liss, Inc.     

Tissue distribution of antihypertensive dipeptide, Val-Tyr, after its single oral administration to spontaneously hypertensive rats.

The distribution of an antihypertensive dipeptide, Val-Tyr (VY), in the tissues of spontaneously hypertensive rats (SHR) was investigated in this study. A single oral administration of VY (10 mg/kg) to 18-week-old SHR resulted in a prolonged reduction of systolic blood pressure (SBP) up to 9 h (SBP0h 198.0+/-3.6 mmHg; SBP9h 154.6+/-3.5 mmHg). As a result of VY determination, a roughly 10-fold higher increment of plasma VY level was observed at 1 h than that at 0 h, whereas thereafter the level declined rapidly. In tissues, VY was widely accumulated in the kidney, lung, heart, mesenteric artery and abdominal aorta with the area under the curve over 9 h of more than 40 pmol h/g tissue; of these a higher VY level was observed in the kidney and lung. In addition, a mean resident time (MRT) for each tissue (>5 h except for liver) revealed that VY preferably accumulated in the tissues rather than in the plasma (MRT 3.8 h). Significant reductions of tissue angiotensin I-converting enzyme activity and angiotensin II level were found in the abdominal aorta as well as in the kidney, suggesting that these organs could be a target site associated with the antihypertensive action of VY.     

Search for substrate-based inhibitors fitting the S2' space of malarial aspartic protease plasmepsin II.

Plasmepsin (Plm) has been identified as an important target for the development of new antimalarial drugs, since its inhibition leads to the starvation of Plasmodium falciparum. A series of substrate-based dipeptide-type Plm II inhibitors containing the hydroxymethylcarbonyl isostere as a transition-state mimic were synthesized. The general design principle was provision of a conformationally restrained hydroxyl group (corresponding to the set residue at the P2' position in native substrates) and a bulky unit to fit the S2' pocket.