Microdissection TESE is superior to conventional TESE in patients with nonobstructive azoospermia caused by Y chromosome microdeletions.

Nonobstructive azoospermia is caused in up to 10% by microdeletions of the Y chromosome in the azoospermia factor (AZF) region, which is divided into three nonoverlapping areas (AZFa, AZFb and AZFc). In 25 male patients with AZF microdeletions, the results of two different techniques for surgical sperm retrieval (SR), conventional multilocular TESE and microdissection TESE, were studied retrospectively over a period of 19 years. Conventional multilocular TESE was carried out in 11 patients and microdissection TESE in 14 patients. Successful SR was possible only in patients with isolated AZFc microdeletions, so only the 20 patients with AZFc microdeletions alone were taken into account for the comparison of the both operative techniques. The sperm detection rate for conventional multilocular TESE was 25%, the sperm detection for microdissection TESE was significantly higher with 67%. In all patients, a histological examination of the testicular tissue was carried out, which showed a mixed picture, but Sertoli-cell-only syndrome in most cases. FSH was no prognostic marker for successful SR. In two of six couples performing an intracytoplasmic sperm injection until now, a pregnancy occurred.

Cross-amplification and characterization of microsatellite loci in Acropora austera from the south-western Indian Ocean.

Here, we report the successful cross-species amplification of previously published acroporid microsatellite markers in the coral Acropora austera from the south-western Indian Ocean. This fast-growing species is a major reef-building coral on South African reefs; however, it is the most damaged coral by scuba diving activity, and is known to be very susceptible to coral bleaching. Neither genetic information nor symbiont-free host tissue was available to develop novel microsatellite markers for this species. Cross-species amplification of previously published microsatellite markers was considered as an alternative to overcome these problems. Of the 21 microsatellite markers tested, 6 were reliably amplified, scored, and found to contain polymorphic loci (3-15 alleles). Although microsatellite sequences are believed to be scarce in the Acropora genome because of its small size, the results of this study and previous research indicate that the microsatellite sequences are well conserved across Acropora species. A detailed screening process identified and quantified the sources of error and bias in the application of these markers (e.g., allele scoring error, failure rates, frequency of null alleles), and may be accounted for in the study of the contemporary gene flow of A. austera in the south-western Indian Ocean.

Reefs and islands of the Chagos Archipelago, Indian Ocean: why it is the world's largest no-take marine protected area.

The Chagos Archipelago was designated a no-take marine protected area (MPA) in 2010; it covers 550 000 km2, with more than 60 000 km2 shallow limestone platform and reefs. This has doubled the global cover of such MPAs.It contains 25-50% of the Indian Ocean reef area remaining in excellent condition, as well as the world's largest contiguous undamaged reef area. It has suffered from warming episodes, but after the most severe mortality event of 1998, coral cover was restored after 10 years.Coral reef fishes are orders of magnitude more abundant than in other Indian Ocean locations, regardless of whether the latter are fished or protected.Coral diseases are extremely low, and no invasive marine species are known.Genetically, Chagos marine species are part of the Western Indian Ocean, and Chagos serves as a 'stepping-stone' in the ocean.The no-take MPA extends to the 200 nm boundary, and. includes 86 unfished seamounts and 243 deep knolls as well as encompassing important pelagic species.On the larger islands, native plants, coconut crabs, bird and turtle colonies were largely destroyed in plantation times, but several smaller islands are in relatively undamaged state.There are now 10 'important bird areas', coconut crab density is high and numbers of green and hawksbill turtles are recovering.Diego Garcia atoll contains a military facility; this atoll contains one Ramsar site and several 'strict nature reserves'. Pollutant monitoring shows it to be the least polluted inhabited atoll in the world. Today, strict environmental regulations are enforced.Shoreline erosion is significant in many places. Its economic cost in the inhabited part of Diego Garcia is very high, but all islands are vulnerable.Chagos is ideally situated for several monitoring programmes, and use is increasingly being made of the archipelago for this purpose.

Male factors determining the outcome of intracytoplasmic sperm injection with epididymal and testicular spermatozoa.

During a period of 8 years, 1,079 intracytoplasmic sperm injection (ICSI) procedures with aspirated epididymal or testicular spermatozoa were performed. Epididymal spermatozoa were used in 172 cycles and testicular spermatozoa or spermatids in 907 cycles. Multiple biopsies were obtained from at least two different locations in the testes. Retrieved spermatozoa were used after cryopreservation (frozen) or immediately after aspiration (fresh). Three hundred patients had obstructive azoospermia (OA) or ejaculation failure. In 414 cases, azoospermia was caused by impaired spermatogenesis resulting from maldescended testes, chemotherapy/radiotherapy, or by Sertoli-cell-only syndrome, genetic disorders or unknown aetiology. Transfer rates, pregnancy rates and birth rates per ICSI cycle showed no statistically significant differences between testicular and epididymal spermatozoa in men with OA (28% average birth rates in both cases). However, birth rates differed significantly with regard to the status of spermatogenesis. Treatment of men with nonobstructive azoospermia (NOA) resulted in a birth rate of 19% per cycle. In all patient groups, there was no difference in the birth rates achieved with fresh and cryopreserved spermatozoa. While testicular volume, follicle-stimulating hormone level and age of the male patient are no statistically significant prognostic factors, the underlying cause of azoospermia is the most important factor determining the outcome of ICSI with epididymal and testicular spermatozoa. The pregnancy rate is lower in NOA patients than in those with OA.

Clathsterol, a novel anti-HIV-1 RT sulfated sterol from the sponge Clathria species.

As part of a search for novel inhibitors of humandeficiency virus type 1 (HIV-1) reverse transcriptase (RT), the MeOH-EtOAc extract of a Red Sea sponge, Clathria sp., was shown to be active. Bioassay-guided fractionation of the extract yielded a novel sterol sulfate, clathsterol (1), which is responsible for the activity and is active at a concentration of 10 microM. The structure of 1 was established mainly by interpretation of spectral data and a chemical transformation.

Cembrane diterpenes from the southern African soft coral Cladiella kashmani.

Two known cembrane diterpenes, flaccidoxide (1) and (1Z,3E,7E,11S, 12S,14S)-11,12-epoxy cembra-1,3,7-trien-14-ol (2), and the new cembrane diterpene flaccidoxide-13-acetate (3) were isolated from specimens of Cladiella kashmani collected off Ponto do Oura, Mozambique. The modified Mosher's method established the previously unassigned absolute configuration of 1 as (1Z,3E,7E,11S,12S,13S, 14R)-14-acetoxy-11,12-epoxy cembra-1,3,7-trien-13-ol. Acetylation of 1 yielded 3 and thus confirmed the structure of 3 as (1Z,3E,7E,11S, 12R,13S,14R)-13,14-diacetoxy-11,12-epoxy cembra-1,3,7-triene. All three diterpenes were toxic to the brine shrimp Artemia salina.

Clathculins A and B, two novel nitrogen-containing metabolites from the sponge Clathrina aff. reticulum.

Two novel acyclic diaza alkenynes, clathculins A and B (1 and 2), were isolated from the Indo-Pacific sponge Clathrina aff. reticulum collected in Sodwana Bay, South Africa. The structure of the mixture of the two unstable inseparable compounds was established by spectroscopic analysis, mainly 1D and 2D NMR measurements and two chemical transformations. Clathculins A and B are the first reported linear marine metabolites containing a 1,2-diaminoethane moiety.

[Fertilization of cryopreserved and thawed human oocytes (Cryo-Oo) by injection of spermatozoa (ICSI)--medical management of sterility and case report of a twin pregnancy]. / Fertilisation von kryokonservierten und aufgetauten humanen Oozyten (Kryo-Oo) vermittels Injektion von Spermatozoen (ICSI)--Sterilitätstherapeutisches Management und Kasuistik einer Zwillingsschwangerschaft.

It is reported on a twin pregnancy and parturition of a healthy girl and a healthy boy following the treatment of the mother by intracytoplasmic injection of spermatozoa (ICSI) into cryopreserved and thawed human oocytes (Cryo-Oo). The couple suffered from a primary sterility because of a severe subfertility of the husband and oligomenorrhea with anovulation of the wife. Before the couple contacted our center for the first time, several pretreatments had been carried out, e.g. intrauterine inseminations (IUI) with husband's and donor sperm and one cycle of in vitro fertilization (IVF) with intracytoplasmic sperm injection. This IVF/ICSI-cycle resulted in a tubal pregnancy. It must be noted that the patient showed in each treatment cycle a remarkable tendency to develop ovarian hyperstimulation syndromes (OHSS), even under stimulation with 50 mg clomiphen citrate. In 1997 we started the first treatment cycle for IVF/ICSI with GnIZH-analogues and FSH according to the long protocol. In spite of a low dosage of gonadotropins the patient again developed a impressive multifollicular growth that we decided, together with the couple, only to perform the oocyte retrieval and cryopreserve the collected eggs. For ethic considerations the couple did not consent in the freezing of pronuclear stages. Two months later we performed the transfer-cycle: after thawing of four oocytes, the intracytoplasmic injection of native spermatozoa led to the fertilization of three oocytes and a total of three preimplantation embryos was transferred one day later. Two weeks later the blood level of hCG was 518 IU/ml and an intact twin pregnancy developed. In July 1998, after 36 weeks of gestation, a healthy girl (2540 g) and an healthy boy (2375 g) were delivered by cesarean section. This case report and the experience with further 23 patients (6 pregnancies) demonstrate that ICSI with cryopreserved and thawed oocytes is an effective approach to avoid repetitive oocyte retrievals. The achievable pregnancy rates so far seem to be similar to frozen pronuclear stages, possibly even better. Under the specific regulations of the German Embryo Protection Act (ESchG)--i.e. freezing of embryos not allowed--this technique is worth being pursued with attention.

Bilosespens A and B: two novel cytotoxic sesterpenes from the marine sponge Dysidea cinerea.

[formula: see text] Two novel sesterpenes, bilosespens A and B (1 and 2) were isolated from the Red Sea sponge Dysidea cinerea collected in the Dahlak archipelago, Eritrea. The structure of the mixture of the two inseparable compounds was established by spectroscopic analysis, mainly by 1D and 2D NMR measurements. The mixture of bilosespens A and B is cytotoxic to a few human cancer cells.

[Intracytoplasmic injection (ICSI) of cryopreserved testicular spermatozoa (Cryo-TESE): a retrospective study of the first 250 treatment cycles]. / Intrazytoplasmatische Injektion (ICSI) von kryokonservierten testikulären Spermatozoen (Kryo-TESE): Eine retrospektive Analyse der ersten 250 Behandlungszyklen.

It is reported on the results of 250 treatment cycles in which we carried out intracytoplasmic injections (ICSI) with frozen and thawed testicular spermatozoa (cryo-TESE). Up to July 1997 we treated 127 patients, 225 embryo transfers were performed (90%), and an average of 2.3 preimplantation embryos were transferred. This resulted in 53 clinical pregnancies, six patients aborted (11.3%). The pregnancy rate was 21.2% per treatment cycle, 23.5% per embryo transfer, and 41.7% per patient. This so called cumulative pregnancy rate is still about to rise, because 49 out of the 72 non-pregnant patients are still in our ICSI-program. Twenty-two children are born, 2 twins and 1 triplet. All children are healthy and without any major malformations. We conclude from these results that using cryopreserved testicular sperm for ICSI is an effective and successful approach for the treatment of severe testicular insufficiency. In comparison to the use of native testicular sperm with the necessity of repetitive testicular biopsies, cryopreservation is advantageous in many concerns (e.g. logistic, organisatoric and financial) and is therefore recommended for clinical routine.

[Intracytoplasmic injection of epididymal and testicular sperm after failed heterologous insemination]. / Zur intrazytoplasmatischen Injektion von epididymalen und testikulären Spermien nach frustranen heterologen Inseminationen.

We report on our experiences with intracytoplasmic injection (ICSI) of epididymal and testicular spermatozoa (MESA, TESE) from azoospermic men whose wives had previously failed to become pregnant after several cycles of artificial insemination by donor (AID); because we do not perform AID treatment in our clinic, all these treatments were carried out in other fertility centers as well as the female diagnostic of sterility. In 3 husbands we could not find any testicular spermatozoa or spermatids, leaving 15 women under treatment. Of these 15 women, 9 became pregnant. This accounts for a pregnancy rate per patient of 60%. We believe that functional defects of the oocytes and somatizing psychological problems concerning AID are predominantly responsible for these results and that both problems can be overcome by ICSI. Besides, these results demonstrate that ICSI/MESA and ICSI/TESE are effective approaches in the treatment of azoospermic men and that using cryopreserved spermatozoa is not disadvantageous in the outcome of ICSI.

Haliclorensin, a Novel Diamino Alkaloid from the Marine Sponge Haliclonatulearensis

Haliclorensin (1), a novel diamino alkaloid possessing an azacyclodecane ring, has been isolated from the sponge Haliclona tulearensis. Its structure was elucidated on the basis of spectroscopic data, as well as by comparison with gamma-amino azacycloalkanes.

New diterpenes from the South African soft coral Eleutherobia aurea.

The known diterpenoids zahavin A and 9-deacetoxy-14,15-deepoxyxeniculin and the two new diterpenoids 7,8-epoxyzahavin A and xeniolide C were isolated from specimens of Eleutherobia aurea collected from Aliwal Shoal off the southern Kwazulu-Natal coast, South Africa. Standard spectroscopic methods were used for the structure determinations. The former three diterpenoids inhibit superoxide production in rabbit-cell neutrophils.

Dizygotic twin pregnancy after intracytoplasmic sperm injection of 1 day old unfertilized oocytes.

We report on a case where late intracytoplasmic sperm injection (ICSI) on unfertilized oocytes after standard in-vitro fertilization (IVF) cycles resulted in a dizygotic twin pregnancy. Fifteen oocytes were harvested from a patient with a history of salpingotomy. After a single cycle of IVF, only one oocyte showed two pronuclei. Subsequently ICSI was performed on six unfertilized metaphase II oocytes, and three of these oocytes showed two pronuclei. Three fertilized embryos were transferred (two derived from ICSI and one from IVF). A normal twin pregnancy resulted, and after delivery of two healthy boys the twins were confirmed to be dizygotic by DNA analysis of several loci. We conclude that at least one of the embryos was derived from the reinsemination by 'second day ICSI'.

Relationship of bacteriological characteristics to semen indices and its influence on fertilization and pregnancy rates after IVF.

BACKGROUND: To determine if routine semen culture is useful in asymptomatic couples joining an in-vitro fertilization (IVF/ET) program. METHODS: Bacterial cultures and semen analysis according to WHO recommendations were performed on semen samples obtained before oocyte recovery from 88 asymptomatic couples undergoing IVF during a 7-month period. RESULTS: In 46 cultures at least one kind of microorganisms could be isolated. Forty-two cultures either contained bacterias regarded as normal skin flora (n = 14) or showed no growth of microorganisms (n = 28). No differences were found in sperm concentration, total sperm count and sperm morphology between the semen samples with positive bacteriology and those with negative culture results. Sperm motility was decreased before Percoll preparation if microorganisms were present. Positive culture results had no effect on either fertilization or pregnancy rates. CONCLUSIONS: These observations suggest that bacteriospermia is not associated with abnormal sperm function after Percoll preparation or adverse IVF outcome.

Evaluation of a cut-off value for normal sperm morphology using strict criteria to predict fertilization after conventional in-vitro fertilization and embryo transfer in asthenozoospermia.

The predictive value of sperm morphology evaluation using strict criteria (MEUSC) on fertilization and pregnancy rates was evaluated in 60 couples undergoing in-vitro fertilization (IVF) and embryo transfer. In all, 47 of the male partners had a progressive sperm motility < 50% with a density > 20 x 10(6)/ml (asthenozoospermia) and 13 had normozoospermia. MEUSC was performed on the same semen sample that was used for IVF on at least 100 spermatozoa after Papanicolaou staining at x 1250 magnification. Defects in the head, mid-piece and tail were counted separately, borderline forms were classified as abnormal and the teratozoospermia index was calculated for each sample. Of the spermatozoa in the asthenozoospermic group, 13.0% were judged as normal on MEUSC versus 19.8% in normozoospermia (P = 0.0013). The number of normal spermatozoa on MEUSC correlated with the progressive motility in asthenozoospermia (rho = 0.41, P = 0.0043). Defects in the mid-piece (P = 0.0004) and tail (P = 0.025) were more common, and the teratozoospermia index (P = 0.015) was higher in asthenozoospermic than normozoospermic samples. The parameters of MEUSC did not correlate with the fertilization rate. In asthenozoospermia, differences in fertilization rates were calculated for each cut-off value between 4 and 10% normal spermatozoa on MEUSC. A cut-off value of 6% gave the best statistical power. If > 6% of spermatozoa were normal, a median of 66.7% of ova were fertilized, compared with a median fertilization rate of 35.4% if < or = 6% of spermatozoa were normal (P = 0.022). The highest cut-off value still giving significant discrimination was 8% (normal MEUSC > 8%, median fertilization rate 66.7%; < or = 8%, median fertilization rate 35.7%, P = 0.028), and this may be used as the critical value for normal morphology in asthenozoospermia. It is concluded that despite a significant linear relationship between morphology using strict criteria and progressive motility, MEUSC is still useful as a predictor for fertilization in asthenozoospermia. In cases of asthenozoospermia with < or = 8% normal spermatozoa on MEUSC, micromanipulation should be considered.

Sodwanones G, H, and I, new cytotoxic triterpenes from a marine sponge.

Three cytotoxic triterpenes, sodwanones G [5], H [6], and I [7], have been isolated from a marine sponge. The three structures were determined by interpretation of nmr spectra, and in the case of 5, also on the basis of the X-ray diffraction analysis. The X-ray diffraction analysis of two other earlier reported sodwanones, E [3] and F [4] is also discussed. The cytotoxic activity against several cancer cell lines, has been tested. IC50 values between 20 and 0.02 microM were obtained. Compounds 5 and 6 were 10- and 500-fold, respectively, more cytotoxic to A-549 cells than to any of the other cell lines tested (HT-29, MEL-28, P-388).

Purification, properties and structural aspects of a thermoacidophilic alpha-amylase from Alicyclobacillus acidocaldarius atcc 27009. Insight into acidostability of proteins.

The alpha-amylase from the thermoacidophilic eubacterium Alicyclobacillus (Bacillus) acidocaldarius strain ATCC 27009 was studied as an example of an acidophilic protein. The enzyme was purified from the culture fluid. On an SDS/polyacrylamide gel, the protein an apparent molecular mass of 160 kDa, which is approximately 15% higher than that predicted from the nucleotide sequence. The difference is due to the enzyme being a glycoprotein. Deglycosylation or synthesis of the enzyme in Escherichia coli gave a product with the mass expected for the mature protein. The amylase hydrolyzed starch at random and from the inside, and its main hydrolysis products were maltotriose and maltose. It also formed glucose from starch (by hydrolysing the intermediate product maltotetraose to glucose and maltotriose) and exhibited some pullulanase activity. the pH and temperature optima were pH3 and 75 degrees C, respectively, characterizing the enzyme as being thermoacidophilic. Alignment of the sequence of the enzyme with that of its closest neutrophilic relatives and with that of alpha-1,4 or alpha-1,6 glycosidic-bond hydrolyzing enzymes of known three-dimensional structure showed that the acidophilic alpha-amylase contains approximately 30% less charged residues than do its closest relatives, that these residues are replaced by neutral polar residues, and that hot spots for these exchanges are likely to be located at the surface of the protein. Literature data show that similar effects are observed in three other acidophilic proteins. It is proposed that these proteins have adapted to the acidic environment by reducing the density of both positive and negative charges at their surface, that this effect circumvents electrostatic repulsion of charged groups at low pH, and thereby contributes to the acidostability of these proteins.

Amino acid derivatives from the marine sponge Jaspis digonoxea.

Six heterocycles, bengamide A [1], bengamide B [2], cyclo(L-trans-(4-hydroxyprolinyl)-L-phenylalanine) [4], the functionalized nonene lactone 5, the novel digonazole [6] and cyclo(L-prolinyl-L-tyrosine) [7], previously unreported from marine origin have been isolated from the South African sponge Jaspis digonoxea. The structures of the known compounds and the new digonazole [6] were elucidated primarily by nmr spectroscopy.

Postpartum blues: relationship between not-protein bound steroid hormones in plasma and postpartum mood changes.

The relationship between non-bound steroid hormone levels in plasma and the occurrence of postpartum mood changes was investigated in 26 newly delivered mothers throughout the first 5 days postpartum. Studies with saliva samples had reported higher concentrations of 17 beta-estradiol and progesterone on the days of symptoms in women experiencing postpartum blues. As there had been a controversy as to how far saliva concentrations reflect free hormone levels in plasma, free hormone levels of 17 beta-estradiol and progesterone were determined in plasma using ultrafiltration. No significant difference concerning free hormone levels could be found between women with and without postpartum blues.