Similar morphokinetic patterns in embryos derived from obese and normoweight infertile women: a time-lapse study.

STUDY QUESTION: Does female obesity affect the dynamic parameters of embryo quality assessed by time-lapse analysis? SUMMARY ANSWER: Female obesity does not affect the dynamic embryo quality as determined by image acquisition and time-lapse analysis. WHAT IS KNOWN ALREADY: Female obesity impairs natural and assisted reproduction but there is no agreement on the specific contribution of gametes, embryos or endometrial receptivity. In this preliminary study the dynamic parameters of embryo quality are assessed for the first time by time-lapse analysis. STUDY DESIGN, SIZE, DURATION: Two-year cohort retrospective study comparing embryos from three groups of patients according to the presence of infertility and/or obesity. PARTICIPANTS AND SETTING: Participants attended a University-affiliated private clinic where ICSI was performed. Using an IVF incubator with a built-in camera designed to automatically acquire images at defined time points, we monitored individual embryos from 89 patients: 71 embryos from 13 obese infertile women, 242 embryos from 45 normoweight infertile women and 111 embryos from 31 normoweight fertile oocyte donors. The chronological pattern of cell divisions (timings of cell cleavages) and other morphologic features (time-dependent cell size and nucleation) was recorded. MAIN RESULTS AND THE ROLE OF CHANCE: Embryos from obese and normoweight infertile women showed similar cleavage patterns, but occurring more slowly, to those from fertile donors. These differences were statistically significant for t2 (time of cleavage to two-blastomere embryo) (P = 0.016), t3 (P = 0.014), t4 (P = 0.003) and t5 (P = 0.040). LIMITATIONS, REASONS FOR CAUTION: These are preliminary data from a retrospective analysis with a limited sample size. GENERALIZABILITY TO OTHER POPULATIONS: Not recommended until further studies using time-lapse analysis of a larger sample have been performed. STUDY FUNDING/COMPETING INTEREST(S): None.

Mosaic results after preimplantation genetic testing for aneuploidy may be accompanied by changes in global gene expression.

Aneuploidy in preimplantation embryos is a major cause of human reproductive failure. Unlike uniformly aneuploid embryos, embryos diagnosed as diploid-aneuploid mosaics after preimplantation genetic testing for aneuploidy (PGT-A) can develop into healthy infants. However, the reason why these embryos achieve full reproductive competence needs further research. Current RNA sequencing techniques allow for the investigation of the human preimplantation transcriptome, providing new insights into the molecular mechanisms of embryo development. In this prospective study, using euploid embryo gene expression as a control, we compared the transcriptome profiles of inner cell mass and trophectoderm samples from blastocysts with different levels of chromosomal mosaicism. A total of 25 samples were analyzed from 14 blastocysts with previous PGT-A diagnosis, including five low-level mosaic embryos and four high-level mosaic embryos. Global gene expression profiles visualized in cluster heatmaps were correlated with the original PGT-A diagnosis. In addition, gene expression distance based on the number of differentially expressed genes increased with the mosaic level, compared to euploid controls. Pathways involving apoptosis, mitosis, protein degradation, metabolism, and mitochondrial energy production were among the most deregulated within mosaic embryos. Retrospective analysis of the duration of blastomere cell cycles in mosaic embryos revealed several mitotic delays compared to euploid controls, providing additional evidence of the mosaic status. Overall, these findings suggest that embryos with mosaic results are not simply a misdiagnosis by-product, but may also have a genuine molecular identity that is compatible with their reproductive potential.

Hand infections.

The hand is an extremely versatile organ adapted for fine tasks with various clinicoanatomical compartments. This article reviews the types of common hand infections that present to the emergency department and/or hand surgeon, with relevant investigations and strategies for diagnosis and treatment, with the emphasis on distinguishing between superficial and more serious infections.

Large outbreak of salmonella phage type 1 infection with high infection rate and severe illness associated with fast food premises.

OBJECTIVES: To describe the epidemiology of an outbreak of Salmonella enteritidis phage type 1 (PT1) infection associated with a fast food premises, and to identify the causative factors leading to an acute outbreak with high attack rate and severe illness including hospital admission. STUDY DESIGN: Integrated descriptive study of epidemiology, food and environmental microbiology, and professional environmental health assessment, supplemented by a case-case analytical study. METHODS: Cases were identified through multiple sources and were interviewed to identify food items consumed. Descriptive epidemiology of all cases and a case-case analytical study of risk factors for severe illness were undertaken. Microbiological investigation included analysis and typing of pathogens from stools, blood and environmental surfaces. Professional environmental heath assessment of the premises was undertaken. RESULTS: S. enteritidis PT1 was recovered from two-thirds of faecal samples. Three cases had dual infection with enterotoxin-producing Clostridium perfringens. S. enteritidis PT1 was isolated from 14 of 40 food samples examined and C. perfringens was isolated from eight food samples. Environmental health inspection of the premises revealed multiple deficiencies, including deficits in food preparation and hygiene consistent with multiple cross-contamination, and time-temperature abuse of sauces widely used across menu items. Severe cases were associated with consumption of chips and salad. CONCLUSIONS: Outbreaks from fast food premises have been infrequently described. This outbreak demonstrates the potential for fast food premises, with multiple deficiencies in food preparation and hygiene, to produce large, intense community outbreaks with high attack rates and severe illness, highly confined in space and time.

Androgen receptor gene CAG trinucleotide repeats in anovulatory infertility and polycystic ovaries.

Hyperandrogenism is currently thought to be central to the pathogenesis of polycystic ovarian syndrome (PCOS), a common endocrine disorder in premenopausal women characterized by irregular menstruation and anovulatory infertility. Although hyperandrogenism is characteristic, some women with PCOS have normal serum androgen levels. All androgens act through the X-linked androgen receptor (AR), the N-terminal domain of which contains a polyglutamine tract encoded by a highly polymorphic CAG trinucleotide repeat tract. Recently, variations in this CAG microsatellite tract, while remaining within the normal polymorphic range (11-38 CAGs), have been inversely correlated with receptor activity. Thus, short tracts are associated with high intrinsic AR activity and increased severity and earlier age of onset of the androgen-regulated tumor prostate cancer, whereas longer CAG tracts are associated with low AR activity and oligospermic infertility. To investigate the role of the CAG repeat tract in PCOS, we measured its length in 91 patients with ultrasound diagnosis of polycystic ovaries, irregular menstrual cycles, and anovulatory infertility and compared them to 112 control subjects of proven fertility with regular menses. Fluorescent-labeled DNA fragments containing the CAG repeat tract were amplified from leucocytic DNA, and their lengths were compared with internal size markers on an automated DNA Sequencer. There were no differences in the mean CAG length between patients and controls when both alleles were considered together or separately. Because there is a subset of PCOS patients whose serum androgens are normal, we compared differences in CAG length between patients whose serum testosterone (T) levels were below the normal laboratory mean, to those that were higher. There was a trend for a lower mean CAG biallelic length among anovulatory patients with T less than 1.73 nmol/L compared with those whose T was more than 1.73 nmol/L (22.47 +/- 0.36 vs. 23.25 +/- 0.29). This difference in CAG length between patients with low and high T levels (20.38 +/- 0.51 vs. 21.98 +/- 0.29) was highly significant (P = 0.004) when only the shorter allele of each individual was considered. Ethnic differences were also evident in our data; Indian subjects had a significantly shorter AR-CAG length compared with Chinese, being 22.08 +/- 0.50 and 23.16 +/- 0.17, respectively. Our data indicate an association between short CAG repeat length and the subset of anovulatory patients with low serum androgens, suggesting that the pathogenic mechanism of polycystic ovaries in these patients could be due to the increased intrinsic androgenic activity associated with short AR alleles.

Internal audit in a microbiology laboratory.

AIM: To set up a programme of internal laboratory audit in a medical microbiology laboratory. METHODS: A model of laboratory based process audit is described. Laboratory activities were examined in turn by specimen type. Standards were set using laboratory standard operating procedures; practice was observed using a purpose designed questionnaire and the data were analysed by computer; performance was assessed at laboratory audit meetings; and the audit circle was closed by re-auditing topics after an interval. RESULTS: Improvements in performance scores (objective measures) and in staff morale (subjective impression) were observed. CONCLUSIONS: This model of process audit could be applied, with amendments to take local practice into account, in any microbiology laboratory.

Trinucleotide (CAG) repeat polymorphisms in the androgen receptor gene: molecular markers of risk for male infertility.

OBJECTIVE: To determine whether changes in the polymorphic trinucleotide (CAG) tract of the androgen receptor gene are associated with spermatogenic defects in patients with male infertility. DESIGN: Case-control study of two ethnic groups. SETTING: University referral centers for male infertility at Baylor College of Medicine, Houston, Texas, and National University Hospital, Singapore. PARTICIPANT(S): Two hundred and fifteen patients with male infertility and depressed spermatogenesis and 142 fertile controls. MAIN OUTCOME MEASURE(S): Size of androgen receptor CAG alleles according to fluorescent-labeled polymerase chain reaction and automated analysis using Genescan software (PE Biosystems Asia, Singapore), and statistical examination of its relation to clinical variables. RESULT(S): In U.S. patients, the mean androgen receptor CAG length was significantly longer in infertile patients than in fertile controls (21.95 +/- 0.31 vs. 20.72 +/- 0.52). Logistic regression showed that each unit increase in CAG length was associated with a 20% increase in the odds of being azoospermic. The odds ratio for azoospermia was sevenfold higher for patients with > or =26 CAG repeats than in those with <26 CAG repeats. Although mean CAG length in Singapore patients was longer than in the U.S. samples, long androgen receptor CAG alleles were significantly related to male infertility in both populations. CONCLUSION(S): Long (> or =26) androgen receptor CAG alleles, which are found in up to 25% of azoospermic men, are associated with male infertility and defective spermatogenesis. Conception in these men is possible with assisted reproductive technologies, as many have spermatozoa in their testes.

Epidemiologically related and unrelated strains of Pseudomonas aeruginosa serotype O12 cannot be distinguished by phenotypic and genotypic typing.

A clonal origin for European isolates of antibiotic multi-resistant Pseudomonas aeruginosa serotype O12 has been suggested. This study was designed to assess the value and limitations of several typing methods for the investigation of outbreaks due to this serotype. In Hôpital de Rodez, France, this organism is endemic, and a prospective clinical epidemiological study was undertaken over a 15 month period, encompassing all patients at the hospital from whom P. aeruginosa O12 was isolated. All isolates were examined by auxanogram, antibiogram, phage-typing, electrophoresis of esterases and pulsed-field gel electrophoresis of DNA. The results suggest that (1) the methods used did not clearly differentiate between clinically-related and epidemiologically-unrelated European isolates, (2) in Hôpital de Rodez, while some isolates were likely to have been transmitted from patient-to-patient, most infections or colonizations with this organism were sporadic and their origin is unknown. The limits of typing methods for the investigation of outbreaks of nosocomial infection with multi-resistant P. aeruginosa O12 are emphasized.

Ribotyping and DNA macrorestriction analysis of isolates of Burkholderia pseudomallei from cases of melioidosis in Malaysia.

Forty-nine isolates of Burkholderia pseudomallei from sporadic cases of melioidosis in Malaysia over the past 18 years were examined by BamHI ribotyping and pulsed-field gel electrophoresis (PFGE) of XbaI digests of total deoxyribonucleic acid (DNA). Twenty-four patients had septicaemic melioidosis with a mortality of 70%; mortality in the non-septicaemic disease was 16%. Five ribotype patterns were identified, 2 of which accounted for 90% of all isolates. PFGE revealed a number of different strains within these ribotypes, but some pairs of isolates from unrelated cases gave closely similar DNA profiles. These results are in agreement with Australian studies which showed a high prevalence of a few ribotypes of B. pseudomallei which are further divisible by genotyping, in areas where melioidosis is endemic.

Drug-related recurrent meningitis.

A 56-year-old man presented with recurrent smear and culture-negative meningitis having ingested Ibuprofen before each episode. The association between Ibuprofen and meningitis has been well established in systemic lupus erythematosus but has been reported only rarely in previously healthy patients.

A preliminary evaluation of a prototype western blot confirmatory test kit for syphilis.

A prototype Western blot kit was evaluated as a confirmatory test for syphilis using 131 sera characterized by other serological tests for syphilis. There were 114 treponemal sera (including 94 cases of early syphilis, 83 of which were untreated) and 17 non-treponemal problem sera (11 gave false positive reactions on screening with the TmpA recombinant antigen enzyme immunoassay (EIA), 3 gave false positive fluorescent treponemal antibody absorbed (FTA-abs) tests, and 3 false positive Captia Syphilis G EIA results). Based on the manufacturer's criteria of reactivity in multiple bands for designating a positive result the Western blot test gave a sensitivity of 99.1% (113/114) and a specificity of 88.2% (15/17) when indeterminate reactions were scored positive and 98.2% (112/114) and 100% (17/17) when indeterminate reactions were scored negative. Sensitivity was high in both treated and untreated infection. Corresponding sensitivities for the TPHA and FTA-abs when equivocal reactions were scored negative were 97.5% (111/114) and 99.1% (113/114). The high sensitivity of the FTA-abs in this study is probably due to the large number of untreated primary infections. Our results with the Western blot, confirm earlier studies using 'in-house' test systems and, support a role for a commercial Western blot test in the confirmatory diagnosis of syphilis. Further studies are required to confirm the high specificity and sensitivity of the kit in a larger series including a wider variety of non-treponemal cases as well as patients with untreated and treated infection.

Genetics of male infertility: role of androgen receptor mutations and Y-microdeletions.

INTRODUCTION: Although infertility affects about 5% of the male population, its cause in most cases is uncertain. Normal spermatogenesis depends on a sequential cascade of genetic events triggered by factors encoded by the sex chromosomes. To determine the contribution of genetic aberrations to male infertility, the X-linked androgen receptor gene and the Y-chromosome were examined for mutations in a large cohort of infertile men. METHODS: Screening of the androgen receptor (AR) gene for single-stranded conformation polymorphisms, confirmation by DNA sequencing, structure-function studies with androgen-responsive reporter genes and chimeric-protein constructs were performed. Y-chromosome microdeletions screening was done with multiplex polymerase chain reaction (PCR) analyses. RESULTS: Genetic screening of over 400 patients and controls showed that defects in the androgen receptor gene lead to the production of dysfunctional receptor protein in 15% of males with abnormally low sperm production. The dozens of mutations and polymorphisms uncovered were associated with reduced intrinsic androgen receptor activity and involve principally two regions of the androgen receptor. Gene-transfer experiments implicated defective intermolecular protein-protein interactions with coactivator molecules as the cause of reduced receptor function. Submicroscopic deletions of the Y-chromosome were also been detected in about 6% of patients with severely reduced spermatogenesis. The deleted segments encoded RNA-binding proteins of unknown function and are not linked to defects in the androgen receptor. CONCLUSIONS: Mutations and polymorphisms of the AR, and Y-microdeletions cause defective sperm production and male infertility in about 20% of subfertile men. These traits can be transmitted to progeny, and counselling can be offered to affected families. Clarification of the molecular mechanisms of pathogenesis has led to rational hormonal therapy.

A laboratory experiment for determining both the hydraulic and diffusive properties and the initial pore-water composition of an argillaceous rock sample: a test with the Opalinus clay (Mont Terri, Switzerland).

Argillaceous formations are thought to be suitable natural barriers to the release of radionuclides from a radioactive waste repository. However, the safety assessment of a waste repository hosted by an argillaceous rock requires knowledge of several properties of the host rock such as the hydraulic conductivity, diffusion properties and the pore water composition. This paper presents an experimental design that allows the determination of these three types of parameters on the same cylindrical rock sample. The reliability of this method was evaluated using a core sample from a well-investigated indurated argillaceous formation, the Opalinus Clay from the Mont Terri Underground Research Laboratory (URL) (Switzerland). In this test, deuterium- and oxygen-18-depleted water, bromide and caesium were injected as tracer pulses in a reservoir drilled in the centre of a cylindrical core sample. The evolution of these tracers was monitored by means of samplers included in a circulation circuit for a period of 204 days. Then, a hydraulic test (pulse-test type) was performed. Finally, the core sample was dismantled and analysed to determine tracer profiles. Diffusion parameters determined for the four tracers are consistent with those previously obtained from laboratory through-diffusion and in-situ diffusion experiments. The reconstructed initial pore-water composition (chloride and water stable-isotope concentrations) was also consistent with those previously reported. In addition, the hydraulic test led to an estimate of hydraulic conductivity in good agreement with that obtained from in-situ tests.

Early onset of progressive subaortic stenosis after complete repair of tetralogy of Fallot.

Tetralogy of Fallot is often found in association with a wide variety of other cardiac lesions, but is rarely found in association with lesions causing obstruction to the left ventricular inflow or outflow. Subaortic stenosis has only rarely been reported in association with tetralogy of Fallot. We report a patient with Marfan syndrome who underwent repair of tetralogy of Fallot at five years of age. Discrete and progressive subaortic stenosis developed two years after the surgical correction, in a previously normal and unobstructed left ventricular outflow tract. Surgical removal of the acquired fibrous subaortic shelf was successful. Clinical signs of obstruction within the left ventricular outflow tract after surgical repair of tetralogy of Fallot should prompt further investigation to exclude the onset of acquired subaortic stenosis.