Killer whale fecal samples: How to get the most out of a single extraction.

Fecal samples are a non-invasive and relatively accessible matrix for investigating physiological processes in resident killer whale (Orcinus orca) populations. The high lipid content of the diet (primarily salmonids) leads to lower density fecal material and slower dispersion, facilitating sample collection. As fecal discharge is relatively infrequent and the volume of sample is variable, maximizing analytical options is an important consideration. Here we present an extraction methodology to measure hormones and lipid content from the same fecal aliquot. Lipid extractions are commonly conducted using chloroform and methanol from Folch or Bligh and Dyer (B&D), while alcohol is the primary solvent for hormone extraction. We evaluated the possibility of using the methanol layer from lipid extractions to assess fecal steroid hormone levels. Folch and B&D methanol residues were assayed form metabolites of progesterone (PMs) and corticosterone (GCs), and results were compared to aliquots extracted in 70 % ethanol. Hormone concentrations measured in the methanol layer from Folch and B&D extractions were 55 % to 79 % lower than concentrations in 70 % ethanol. We developed mathematical corrections, using linear regression models fitted to Folch or B&D methanol vs 70 % ethanol hormone concentrations (p < 0.01). Fecal concentrations of PMs and GCs from methanol extractions were biologically validated and are significantly higher in confirmed pregnant females compared to non-pregnant individuals (p < 0.05). This study demonstrates that lipid extraction protocols may be used for the analysis of multiple biomarkers, maximizing the use of small-volume samples.

Historical and contemporary perspectives on children's diets: is choice always in the patients' best interest?

On 29 March 1744, Thomasin Grace, a 13-year-old girl, was the first inpatient admitted to the Northampton General Infirmary (later the Northampton General Hospital). Inpatient hospital diets, then and now, are mainstays of effective patient treatment. In the mid-18th century there were four prescribed diets at Northampton: 'full', 'milk', 'dry' and 'low'. Previous opinions concerning these four diets were unfavourable, but had not been based upon an individual dietetic assessment. Thomasin would most likely have been given the milk diet, but use of the full diet cannot be excluded. 'Grace Everyman' is Thomasin's modern equivalent. Under current NHS guidelines Thomasin would be considered a paediatric patient, but in 1744 she would have been considered as an adult. This study undertakes a full dietetic analysis of all the prescribed diets available for Thomasin in 1744 and compares this against random choices for Grace from the 2009 inpatient menu from the paediatric (Paddington) ward, and the adult ward inpatient menu at the Northampton General Hospital. The results show that, for Thomasin, the 1744 milk and full diets met the current advised nutritional requirements for adequate dietary intake. However, for Grace, the present 2009 Paddington and adult ward menu, although generally meeting nutritional requirements, could, if Grace or her carer consistently chose poorly during a prolonged inpatient stay, lead to inadequate nutrition. This challenges assumptions that hospital diets were historically inadequate, and that choice in present day equates with satisfactory nutritional intake.

Cystic fibrosis-related diabetes.

Diabetes mellitus (DM) has been recognized as a complication of cystic fibrosis (CF) for almost 50 years and commonly develops around 20 years of age. The prevalence increases with age and, with improved survival of those with CF, approaches 30% in certain centres. Its development appears to have a significant impact on pulmonary function and may increase mortality by up to six-fold. Subjects with CF are rarely ketosis-prone and phenotypically lie between Type 1 and Type 2 DM. Microvascular complications are recognized, although paucity of data does not permit a clear description of their natural history. An annual oral glucose tolerance test from the age of 10 years is recommended for screening, but logistical difficulties have led some groups to develop specific algorithms to aid diagnosis. Insulin sensitivity in CF is much debated and may depend upon the degree of glucose intolerance. Insulin resistance occurs in the presence of infection, corticosteroid usage and hyperglycaemia, whilst hepatic insulin resistance is considered an adaptation to CF. There is no universal consensus on the treatment of hyperglycaemia. With increased longevity of individuals with CF, greater numbers will develop diabetes and the diabetes physician is destined to play a greater role in the multidisciplinary CF team.

Effects of forced diving on the spleen and hepatic sinus in northern elephant seal pups.

In phocid seals, an increase in hematocrit (Hct) accompanies diving and periods of apnea. The variability of phocid Hct suggests that the total red cell mass is not always in circulation, leading researchers to speculate on the means of blood volume partitioning. The histology and disproportionate size of the phocid spleen implicates it as the likely site for RBC storage. We used magnetic resonance imaging on Northern elephant seals to demonstrate a rapid contraction of the spleen and a simultaneous filling of the hepatic sinus during forced dives (P < 0.0001, R(2) = 0.97). The resulting images are clear evidence demonstrating a functional relationship between the spleen and hepatic sinus. The transfer of blood from the spleen to the sinus provides an explanation for the disparity between the timing of diving-induced splenic contraction ( approximately 1-3 min) and the occurrence of peak Hct (15-25 min). Facial immersion was accompanied by an immediate and profound splenic contraction, with no further significant decrease in splenic volume after min 2 (Tukey-Kramer HSD, P = 0.05). At the conclusion of the dive, the spleen had contracted to 16% of its predive volume (mean resting splenic volume = 3,141 ml +/- 68.01 ml; 3.54% of body mass). In the postdive period, the spleen required 18-22 min to achieve resting volume, indicating that this species may not have sufficient time to refill the spleen when routinely diving at sea, which is virtually continuous with interdive surface intervals between 1 and 3 min.

Successful pregnancy and birth after IVF in a woman with cystic fibrosis.

As survival increases, patients with cystic fibrosis (CF) are often confronted with reproductive issues. Initial reports gave conflicting advice regarding the outcome of pregnancy in CF. However a recent large longitudinal study of pregnancies in CF women suggested that pregnancy has little impact on morbidity or mortality. Reduced fertility in CF women has been described, possibly due to thickened cervical mucus, and intrauterine insemination (IUI) has been used to overcome this. We report the first woman with CF, to our knowledge, to be successfully treated with IVF after repeated failed attempts at IUI.

Malnutrition, gastroenteritis and trypsinogen concentration in hospitalised Aboriginal children.

OBJECTIVE: To explore relationships between malnutrition and pancreatic damage in hospitalised aboriginal children. METHODS: Immunoreactive trypsinogen (IRT) concentrations were measured in two populations of hospitalised aboriginal children in Australia: 472 children aged 0-3 years, in Alice Springs (Northern Territory); and 187 children aged 0-16 years in Mount Isa (Queensland). Correlation of whole blood IRT with height and weight z-scores, four-site skinfold thickness and upper arm circumference was sought. RESULTS: In Mount Isa, the geometric mean IRT concentration rose with decreasing weight z-score. The IRT concentration was otherwise unrelated to nutritional indices. Sixty percent of the 39 Mount Isa patients with gastroenteritis and 24.5% of the 358 Alice Springs patients with gastroenteritis had an IRT concentration in the upper quartile for their population, compared with 16% for patients with other diagnoses in both populations. CONCLUSIONS: A high IRT concentration in patients with low weight z-scores is a confounding effect of gastroenteritis, and may result from subclinical pancreatic disease in gastroenteritis.

Increased polymorphonuclear granulocytes in seminal plasma in relation to sperm morphology.

Much controversy surrounds the clinical significance of an increased concentration of white blood cells (WBC) in the male ejaculate. The World Health Organization's classification of leukocytospermia is a concentration > 1 x 10(6) WBC/ml. The aim of this study was to assess the association of varying concentrations of leukocytes to sperm morphology evaluated by strict criteria. Semen samples were collected from a total of 79 patients. Round cells on the initial semen analysis were stained for identification of polymorphonuclear granulocytes (PMN) as the largest group (50-60%) of white blood cells using the Endtz Method commercially produced as Leucoscreen. Diff Quick Staining Kit was used for sperm morphology assessment and 200 spermatozoa were assessed per slide. Data were evaluated using two cut-off criteria, at 0.5 x 10(6) WBC/ml and 1 x 10(6) WBC/ml. Mann-Whitney U-values at both < and > 0.5 x 10(6)/ml PMN (P < 0.001) and at < and > 1.0 x 10(6)/ml PMN (P < 0.015) showed differences between percentage normal forms. Spearman's rank correlation coefficient for PMN concentration showed a negative correlation (P < 0.01) with percentage normal sperm morphology and positive correlation for midpiece abnormalities (P < 0.04). These data support the hypothesis that PMN have a role in the increase of abnormal spermatozoa, particularly those with midpiece abnormalities, by as yet unknown mechanisms.

The effects of clomiphene citrate and cyclofenil on cervical mucus volume and receptivity over the periovulatory period.

OBJECTIVE: To determine the effects of clomiphene citrate (CC) and cyclofenil on cervical mucus (CM) volume and receptivity sampled serially over the periovulatory period. DESIGN: Using prospective luteinizing hormone (LH) timing CM volume and receptivity were compared in standard CC and cyclofenil-stimulated cycles using normal ovulatory cycles as controls. LOCATION: The Donor Insemination Unit at the University Research Clinic, Sheffield, United Kingdom. PATIENTS: Twenty anovulatory patients and 10 normally ovulating patients, all of whom were participating in a treatment cycle of donor insemination. INTERVENTIONS: The 20 anovulatory patients were allocated at random into two groups: group 1 was administered 50 mg of CC on days 2 to 6 of the menstrual cycle; group 2 was administered 400 mg of cyclofenil on days 3 to 12 of the menstrual cycle. All the patients were given a single treatment of donor insemination 24 to 36 hours after the onset of the LH surge. RESULTS: Clomiphene citrate and cyclofenil were shown to exert differential impacts on CM quantity and quality. In terms of quantity, the CC patients produced significantly lower volumes of CM than the cyclofenil patients and controls. In terms of quality, the CC patients and controls produced CM of similar receptivity, whereas the cyclofenil patients produced CM that was significantly more receptive to sperm than both the CC patients and controls. CONCLUSIONS: Neither CC nor cyclofenil exerted a detrimental impact on CM quality throughout the periovulatory period.

Home monitoring of gonadotropin ovulation induction using the Ovarian Monitor.

The safe use of gonadotropins relies on close hormonal and/or ultrasound monitoring to assess the response to treatment, requiring multiple hospital visits. Home monitoring with the Ovarian Monitor (St. Michael Research Foundation, Macleod, Victoria, Australia) minimizes hospital visits and overcomes many of the logistic difficulties associated with gonadotropin use. It utilizes a system of homogenous enzyme immunoassay using lysozyme conjugates to measure quantitatively either urinary estrone-3 or pregnanediol-3-glucuronide. Results obtained by 24 patients in 57 cycles using the Ovarian Monitor at home correlate closely with results obtained in the laboratory (estrone-3-glucuronide r = 0.955; pregnanediol-3-glucuronide r = 0.958). Cycle outcomes (ovulation, 74%/cycle; clinical pregnancy, 30%/cycle; multiple pregnancy, 13%/pregnancy; hyperstimulation, 11%/cycle) are no different from those achieved in laboratory-monitored patients. Home monitoring can be as safe and effective as laboratory monitoring, offers significant social benefits, and improves access to this form of therapy.

Increasing the human menopausal gonadotropin dose--does the response really improve?

This study assesses the effects of attempts to optimize human menopausal gonadotropin (hMG) dosage in 271 patients who had at least two hyperstimulation cycles for in vitro fertilization or gamete intrafallopian transfer. In the first cycle, all patients received clomiphene citrate and hMG 150 IU/d. In the second cycle, the hMG dose was increased in 45% of patients to try to increase the egg yield. In spite of the increase, the population response was practically identical in both cycles. Median numbers of eggs retrieved (6 versus 6), no eggs retrieved (0.4% versus 1%), only one or two eggs retrieved (10% versus 10%), and canceled cycles (10% versus 10.7%). This suggests that increasing the hMG dosage above 150 IU does not increase the number of eggs retrieved. A poor response may be due to inherent differences in follicular development that cannot be overcome by increases in hMG dosage.

Human chorionic gonadotropin to oocyte retrieval interval in in vitro fertilization--how critical is it ?

This study shows that modification of the hCG-oocyte retrieval interval from 34 1/2 hours to 37 hours in patients having IVF had no significant effect on the number or quality of oocytes retrieved or on the ultimate pregnancy rate. More consideration can therefore be given to other factors such as patient convenience when scheduling the timing of hCG administration.