Case Report: Congenital Methaemoglobinaemia Type 1: Benign Cyanosis?

Type 1 congenital methaemoglobinaemia is a rare cause of cyanosis which may manifest in affected individuals during concomitant illness. Treatment indications and aims differ from that of acquired methaemoglobinaemia. Type 1 methaemoglobinaemia is a distinct condition from the type 2 form which has a high mortality rate in infancy. A 25 year old male with known type 1 congential methaemoglobinaemia presented with cyanosis in the context of Influenza A with raised methaemoglobin levels on arterial blood gas analysis. The patient was assessed based on his level of 'functional haemoglobin' with no acute indication for IV methylene blue or ascorbic acid. Consideration could be given to prescription of these on a cosmetic basis for some patient populations.

Discovery of a Ni2+-dependent guanidine hydrolase in bacteria.

Nitrogen availability is a growth-limiting factor in many habitats1, and the global nitrogen cycle involves prokaryotes and eukaryotes competing for this precious resource. Only some bacteria and archaea can fix elementary nitrogen; all other organisms depend on the assimilation of mineral or organic nitrogen. The nitrogen-rich compound guanidine occurs widely in nature2-4, but its utilization is impeded by pronounced resonance stabilization5, and enzymes catalysing hydrolysis of free guanidine have not been identified. Here we describe the arginase family protein GdmH (Sll1077) from Synechocystis sp. PCC 6803 as a Ni2+-dependent guanidine hydrolase. GdmH is highly specific for free guanidine. Its activity depends on two accessory proteins that load Ni2+ instead of the typical Mn2+ ions into the active site. Crystal structures of GdmH show coordination of the dinuclear metal cluster in a geometry typical for arginase family enzymes and allow modelling of the bound substrate. A unique amino-terminal extension and a tryptophan residue narrow the substrate-binding pocket and identify homologous proteins in further cyanobacteria, several other bacterial taxa and heterokont algae as probable guanidine hydrolases. This broad distribution suggests notable ecological relevance of guanidine hydrolysis in aquatic habitats.

The importance of chain context in assessing small nucleotide variants in titin: in silico case study of the I10-I11 tandem and its arrhythmic right ventricular cardiomyopathy linked position T2580.

Non-synonymous small nucleotide variations (nsSNVs) in the giant muscle protein, titin, have key roles in the development of several myopathologies. Although there is considerable motive to screen at-risk individuals for nsSNVs, to identify patients in early disease stages while therapeutic intervention is still possible, the clinical significance of most titin variations remains unclear. Therefore, there is a growing need to establish methods to classify nsSNVs in a simple, economic and rapid manner. Due to its strong correlation to arrhythmogenic right ventricular cardiomyopathy (ARVC), one particular mutation in titin-T2580I, located in the I10 immunoglobulin domain-has received considerable attention. Here, we use the I10-I11 tandem as a case study to explore the possible benefits of considering the titin chain context-i.e. domain interfaces-in the assessment of titin nsSNVs. Specifically, we investigate which exchanges mimic the conformational molecular phenotype of the T2580I mutation at the I10-I11 domain interface. Then, we computed a residue stability landscape for domains alone and in tandem to define a Domain Interface Score (DIS) which identifies several hotspot residues. Our findings suggest that the T2580 position is highly sensitive to exchange and that any variant found in this position should be considered with care. Furthermore, we conclude that the consideration of the higher order structure of the titin chain is important to gain accurate insights into the vulnerability of positions in linker regions and that titin nsSNV prediction benefits from a contextual analysis. Communicated by Ramaswamy H. Sarma.

Influence of fatty acid chain length and unsaturation on mid-infrared milk analysis.

Our first objective was to optimize center wavelengths and bandwidths for virtual filters used in a Fourier transform mid-infrared (MIR) milk analyzer. Optimization was accomplished by adjusting center wavelengths and bandwidths to minimize the size of intercorrection factors. Once optimized, the virtual filters were defined as follows: fat B sample, 3.508 microm (2,851 cm(-1)), and bandwidth of 0.032 microm (26 cm(-1)); fat B reference, 3.556 microm (2812 cm(-1)), and bandwidth of 0.030 microm (24 cm(-1)); lactose sample, 9.542 microm (1,048 cm(-1)), and bandwidth of 0.092 microm (20 cm(-1)); lactose reference, 7.734 microm (1,293 cm(-1)), and bandwidth of 0.084 microm (14 cm(-1)); protein sample, 6.489 microm (1,541 cm(-1)), and bandwidth of 0.085 microm (20 cm(-1)); protein reference, 6.707 microm (1491 cm(-1)), and bandwidth of 0.054 microm (12 cm(-1)); fat A sample, 5.721 microm (1,748 cm(-1)), and bandwidth of 0.052 microm (16 cm(-1)); fat A reference, 5.583 microm (1,791 cm(-1)), and bandwidth of 0.050 microm (16 cm(-1)). The bandwidth and its proximity to areas of intense water absorption had the largest effect on the intercorrection factors. The second objective was to quantify the impact of fatty acid chain length and unsaturation on fat B and fat A MIR measurements. Increasing the chain length increased the difference (i.e., MIR minus reference) between MIR prediction and reference chemistry by 0.0429% and by -0.0566% fat per unit of increase in carbon number per 1% change in fat, for fat B and fat A, respectively. Increasing the unsaturation decreased the difference (i.e., MIR minus reference) between MIR prediction of fat and chemistry for fat B by -0.4021% and increased fat A by 0.0291% fat per unit of increase in double bonds per 1% change in fat concentration.

Impact of fatty acid composition on the accuracy of mid-infrared fat analysis of farm milks.

Our objective was to determine whether data from a previous study using model milk emulsions to characterize the influence of variation in fatty acid chain length and unsaturation on mid-infrared (MIR) fat predictions could be used to identify a strategy to improve the accuracy of MIR fat predictions on a population of farm milks with a wide variation in fatty acid chain length and unsaturation. The mean fatty acid chain length for 45 farm milks was 14.417 carbons, and the mean unsaturation was 0.337 double bonds per fatty acid. The range of fatty acid chain lengths across the 45 farm milks was 1.23 carbons, and the range in unsaturation was 0.167 double bonds per fatty acid. Fat B (absorbance by the carbon-hydrogen stretch) MIR predictions increased and fat A MIR (absorbance by the ester carbonyl stretch) predictions decreased relative to reference chemistry with increasing fatty acid chain length. When the fat B MIR fat predictions were corrected for sample-to-sample variation in unsaturation, the positive correlation between fat B and fatty acid chain length increased from a coefficient of determination of 0.42 to 0.89. A 45:55 ratio of fat B corrected for unsaturation and fat A gave a smaller standard deviation of the difference between MIR prediction and reference chemistry than any ratio of the fat B (without correction for unsaturation) and fat A or either fat B or fat A alone. This demonstrates the technical feasibility of this approach to improve MIR testing accuracy for fat, if a simple procedure could be developed to determine the unsaturation of fat in milk rapidly and to correct the fat B reading for the effect of unsaturation before being combined with fat A.

Lipolysis and proteolysis of modified and producer milks used for calibration of mid-infrared milk analyzers.

Our objective was to determine if lipolysis or proteolysis of calibration sets during shelf life influenced the mid-infrared (MIR) readings or calibration slopes and intercepts. The lipolytic and proteolytic deterioration was measured for 3 modified milk and 3 producer milk calibration sets during storage at 4 degrees C. Modified and producer milk sets were used separately to calibrate an optical filter and virtual filter MIR analyzer. The uncorrected readings and slopes and intercepts of the calibration linear regressions for fat B, fat A, protein, and lactose were determined over 28 d for modified milks and 15 d for producer milks. It was expected that increases in free fatty acid content and decreases in the casein as a percentage of true protein of the calibration milks would have an effect on the MIR uncorrected readings, calibration slopes and intercepts, and MIR predicted readings. However, the influence of lipolysis and proteolysis on uncorrected readings was either not significant, or significant but very small. Likewise, the amount of variation accounted for by day of storage at 4 degrees C of a calibration set on the calibration slopes and intercepts was also very small. Most of the variation in uncorrected readings and calibration slopes and intercepts were due to differences between the optical filter and virtual filter analyzers and differences between the pasteurized modified milk and raw producer milk calibration sets, not due to lipolysis or proteolysis. The combined impact of lipolysis and proteolysis on MIR predicted values was <0.01% in most cases.

Calibration of infrared milk analyzers: modified milk versus producer milk.

Mid-infrared (MIR) milk analyzers are traditionally calibrated using sets of preserved raw individual producer milk samples. The goal of this study was to determine if the use of sets of preserved pasteurized modified milks improved calibration performance of MIR milk analyzers compared with calibration sets of producer milks. The preserved pasteurized modified milk sets exhibited more consistent day-to-day and set-to-set calibration slope and intercept values for all components compared with the preserved raw producer milk calibration sets. Pasteurized modified milk calibration samples achieved smaller confidence interval (CI) around the regression line (i.e., calibration uncertainty). Use of modified milk calibration sets with a larger component range, more even distribution of component concentrations within the ranges, and the lower correlation of fat and protein concentrations than producer milk calibration sets produced a smaller 95% CI for the regression line due to the elimination of moderate and high leverage samples. The CI for the producer calibration sets were about 2 to 12 times greater than the CI for the modified milk calibration sets, depending on the component. Modified milk calibration samples have the potential to produce MIR milk analyzer calibrations that will perform better in validation checks than producer milk-based calibrations by reducing the mean difference and standard deviation of the difference between instrument values and reference chemistry.

Modified versus producer milk calibration: mid-infrared analyzer performance validation.

Our objective was to determine the validation performance of mid-infrared (MIR) milk analyzers, using the traditional fixed-filter approach, when the instruments were calibrated with producer milk calibration samples vs. modified milk calibration samples. Ten MIR analyzers were calibrated using producer milk calibration sample sets, and 9 MIR milk analyzers were calibrated using modified milk sample sets. Three sets of 12 validation milk samples with all-laboratory mean chemistry reference values were tested during a 3-mo period. Calibration of MIR milk analyzers using modified milk increased the accuracy (i.e., better agreement with chemistry) and improved agreement between laboratories on validation milk samples compared with MIR analyzers calibrated with producer milk samples. Calibration of MIR analyzers using modified milk samples reduced overall mean Euclidian distance for all components for all 3 validation sets by at least 24% compared with MIR analyzers calibrated with producer milk sets. Calibration with modified milk sets reduced the average Euclidian distance from all-laboratory mean reference chemistry on validation samples by 40, 25, 36, and 27%, respectively for fat, anhydrous lactose, true protein, and total solids. Between-laboratory agreement was evaluated using reproducibility standard deviation (s(R)). The number of single Grubbs statistical outliers in the validation data was much higher (53 vs. 7) for the instruments calibrated with producer milk than for instruments calibrated with modified milk sets. The s(R) for instruments calibrated with producer milks (with statistical outliers removed) was similar to data collected in recent proficiency studies, whereas the s(R) for instruments calibrated with modified milks was lower than those calibrated with producer milks by 46, 52, 61, and 55%, respectively for fat, anhydrous lactose, true protein, and total solids.

Precalibration evaluation procedures for mid-infrared milk analyzers.

The purpose of this paper is to present a detailed account of the precalibration procedures developed and implemented by the USDA Federal Milk Market Administrators (FMMA) for evaluating mid-infrared (MIR) milk analyzers. Mid-infrared analyzers specifically designed for milk testing provide a rapid and cost-effective means for determining milk composition for payment and dairy herd improvement programs. These instruments determine the fat, protein, and lactose content of milk, and enable the calculation of total solids, solids-not-fat, and other solids. All MIR analyzers are secondary testing instruments that require calibration by chemical reference methods. Precalibration is the process of assuring that the instrument is in good working order (mechanically and electrically) and that the readings before calibration are stable and optimized. The main components of precalibration are evaluation of flow system integrity, homogenization efficiency, water repeatability, zero shift, linearity, primary slope, milk repeatability, purging efficiency, and establishment of intercorrection factors. These are described in detail and apply to both filter-based and Fourier transform infrared instruments operating using classical primary and reference wavelengths. Under the USDA FMMA Precalibration Evaluation Program, the precalibration procedures were applied longitudinally over time using a wide variety of instruments and instrument models. Instruments in this program were maintained to pass the criteria for all precalibration procedures. All instruments used similar primary wavelengths to measure fat, protein, and lactose but there were differences in reference wavelength selection. Intercorrection factors were consistent over time within all instruments and similar among groups of instruments using similar primary and reference wavelengths. However, the magnitude and sign of the intercorrection factors were significantly affected by the choice of reference wavelengths.

Indirect and direct determination of the casein content of milk by Kjeldahl nitrogen analysis: collaborative study.

The classic method for determination of milk casein is based on precipitation of casein at pH 4.6. Precipitated milk casein is removed by filtration and the nitrogen content of either the precipitate (direct casein method) or filtrate (noncasein nitrogen; NCN) is determined by Kjeldahl analysis. For the indirect casein method, milk total nitrogen (TN; Method 991.20) is also determined and casein is calculated as TN minus NCN. Ten laboratories tested 9 pairs of blind duplicate raw milk materials with a casein range of 2.42-3.05% by both the direct and indirect casein methods. Statistical performance expressed in protein equivalents (nitrogen x 6.38) with invalid and outlier data removed was as follows: NCN method (wt%), mean = 0.762, sr = 0.010, sR = 0.016, repeatability relative standard deviation (RSDr) = 1.287%, reproducibility relative standard deviation (RSDr) = 2.146%; indirect casein method (wt%), mean = 2.585, repeatability = 0.015, reproducibility = 0.022, RSDr = 0.560%, RSDR = 0.841; direct casein method (wt%), mean = 2.575, sr = 0.015, sR = 0.025, RSDr = 0.597%, RSDR = 0.988%. Method performance was acceptable and comparable to similar Kjeldahl methods for determining nitrogen content of milk (Methods 991.20, 991.21, 991.22, 991.23). The direct casein, indirect casein, and noncasein nitrogen methods have been adopted by AOAC INTERNATIONAL.

Performance evaluation of direct forced-air total solids and Kjeldahl total nitrogen methods: 1990 through 1995.

Results from collaborative studies of the performance of the direct forced-air oven-drying method for determination of milk total solids content (AOAC Method 990.20) and the Kjeldahl total nitrogen method for determination of milk total nitrogen content (AOAC Method 991.20) were published in 1989 and 1990, respectively. Method performance was characterized by using the harmonized ISO/IU-PAC/AOAC guidelines for method validation, and the methods now have final action status. During 1990 through 1995, the split sample collaborative study format was used to monitor the performance of these methods as part of a multilaboratory quality assurance program. Seven blind duplicate milk materials were sent from a central laboratory once every 2 months to participating laboratories. Data were analyzed with the same statistical procedures used in the original collaborative studies. Compared with the original collaborative study, the repeatability and reproducibility of the oven-drying method improved over time. For the Kjeldahl total nitrogen method, within-laboratory repeatability improved slightly, whereas between-laboratory reproducibility was similar to but not always as good as in the original study. The results demonstrate that the statistical protocol for collaborative studies can be used effectively as the basis for a multilaboratory quality assurance program and that the method performance achieved in a collaborative study can be maintained and even improved with time.

Multilevel anterior cervical corpectomy and fibular allograft fusion for cervical myelopathy.

This study was conducted to determine the safety and efficacy of multilevel anterior cervical corpectomy and stabilization using fibular allograft in patients with cervical myelopathy. Thirty-six patients underwent this procedure for cervical myelopathy caused by spondylosis (20 patients), ossified posterior longitudinal ligament (four patients), trauma (one patient), or a combination of lesions (11 patients). The mean age (+/- standard deviation) of the patients was 58 +/- 10 years and 30 of the patients were men. The mean duration of symptoms before surgery was 30 +/- 6 months and 11 patients had undergone previous surgery. Prior to surgery, the mean Nurick grade of the myelopathy was 3.1 +/- 1.4. Seventeen patients also had cervicobrachial pain. Four vertebrae were removed in six patients, three in 19, and two in 11 patients. Instrumentation was used in 15 cases. The operative mortality rate was 3% (one patient) and two patients died 2 months postoperatively. Postoperative complications included early graft displacement requiring reoperation (three patients), transient dysphagia (two patients), cerebrospinal fluid leak treated by lumbar drainage (three patients), myocardial infarction (two patients), and late graft fracture (one patient). One patient developed transient worsening of myelopathy and three developed new, temporary radiculopathies. All patients achieved stable bone union and the mean Nurick grade at an average of 31 +/- 20 months (range 0-79 months) postoperatively was 2.4 +/- 1.6 (p < 0.05, t-test). Cervicobrachial pain improved in 10 (59%) of the 17 patients who had preoperative pain and myelopathy improved at least one grade in 17 patients (47%; p < 0.05). Twenty-six surviving patients (72%) were followed for more than 24 months and stable, osseous union occurred in 97%. These results show that extensive, multilevel anterior decompression and stabilization using fibular allograft can be achieved with a perioperative mortality and major morbidity rate of 22% and with significant improvement in pain and myelopathy.

Comparison of Babcock and ether extraction methods for determination of fat content of cream: collaborative study.

A modified Mojonnier ether extraction method for determination of the fat content of cream was developed based on the method for milk (AOAC Official Method 989.05). The cream Babcock method (AOAC Official Method 920.111B-C) was modified to harmonize with the milk Babcock method (AOAC Official Method 989.04) and to clarify procedural details. Using the AOAC collaborative study format, 10 laboratories tested 9 pairs of blind duplicate heat-treated cream samples with a fat range of 30-45% using both methods. The statistical performance (invalid and outlier data removed) was as follows: mean % fat = 37.932, Sr = 0.125, SR = 0.151, RSDr = 0.330, RSDR = 0.398, r = 0.354, and R = 0.427 for the ether extraction method. For the Babcock method, mean % fat = 38.209, Sr = 0.209, SR = 0.272, RSDr = 0.548, RSDR = 0.712, r = 0.592, and R = 0.769. Average test results for fat from the Babcock method were 0.277% (absolute fat) greater than for the Mojonnier ether extraction method. The difference between methods, as a percentage of the average fat content of the samples, was 0.73%. This agrees with differences observed between the 2 methods for milk when 10 to 17 laboratories tested 7 milk samples in blind duplicate at bimonthly intervals over a 4-year period (average difference 0.029% fat, 0.78% as a percentage of average fat content). The Mojonnier ether extraction and Babcock methods for fat in cream have been adopted by AOAC INTERNATIONAL. The new Babcock method replaced the AOAC Official Method 920.111B-C.

Effect of infrared analyzer homogenization efficiency on repeatability of uncorrected fat A and fat B signals.

Poor repeatability by infrared milk analyzers may be caused by inefficient homogenization as a result of light scattering and the Christiansen effect. The objectives of this study were to identify instruments with good and poor homogenization efficiency and to determine if a difference exists in repeatability performance between instruments with good vs poor homogenization efficiency. Unhomogenized and homogenized portions of the same milk were tested 20 times consecutively on 22 instruments. An instrument was considered to have poor homogenization efficiency if the mean difference in the uncorrected signal between unhomogenized and homogenized portions of the same milk was > or = 1.43% of the fat test (i.e., > or = 0.05% at 3.5% fat). Instruments were evaluated for repeatability by calculating the sample standard deviation and the range of the latter 19 uncorrected readings for unhomogenized and homogenized milks. When repeatability was evaluated as a function of homogenization efficiency, there was a significant (p = 0.001) correlation between poor homogenization efficiency and poor repeatability when testing unhomogenized milk but not when testing homogenized milk. Improved homogenizer performance within infrared milk analyzers is needed to improve the repeatability of raw milk testing.

What in the world is being done about TBAs? An overview of international and national attitudes to traditional birth attendants.

Many of the half million women per year who die in childbirth are attended by traditional birth attendants (TBAs). Whether they fare better when such an attendant is trained remains uncertain; even the World Health Organization seems to have tempered its enthusiasm for TBA training recently. With some nations outlawing the practice of TBAs and others actively promoting it, there seems to be no consensus on what to do about this major and continuing workforce in maternity care. By themselves TBAs cannot reduce maternal mortality, whether they are trained or not. They need skilled, equipped and available support. As the professional group who must co-operate with TBAs and provide that support, midwives must, collectively and individually, assess, state and act on their attitude towards TBAs.

Effect of infrared analyzer homogenization efficiency on linearity of uncorrected fat A and fat B signals.

The objective of the survey was to determine if poor homogenizer performance causes nonlinear behavior of the uncorrected fat A or fat B signal that is not detected when an instrument's residual nonlinearity is determined by using dilutions of homogenized milk instead of unhomogenized milk. Unhomogenized and homogenized (17238 kPa) portions of the same 6.1% fat milk were tested on 20 instruments to determine homogenization efficiency. Instruments with differences of > or = 0.087% fat between homogenized and unhomogenized portions of the same milk had inefficient homogenization, on the basis of criteria established in a previous study. Four and 12 instruments out of 20 demonstrated inefficient homogenization for the fat A and fat B channels, respectively. Uncorrected signal linearity for the fat channels was evaluated quantitatively by using a series of dilutions of homogenized (17238 kPa) and unhomogenized milks. Most instruments passed the linearity evaluation for dilutions of either homogenized or unhomogenized milk, even though many of the same instruments failed the homogenization efficiency evaluation. Thus, using dilutions of homogenized milk is valid for linearity evaluation of instruments being used for testing unhomogenized milk in the range of fat concentrations used for payment testing.

Induction of lactation: histological and biochemical development of mammary tissue and milk yields of cows injected with estradiol-17 beta and progesterone for 21 days.

Lactations were induced in nonpregnant, nonlactating dairy cows by subcutaneous injections of estradiol-17 beta and progesterone for 21 d (.10 and .25 mg/kg body weight/d) and dexamethasone (.028 mg/kg body weight/d) on d 31 to 34. Milking was initiated on d 35. Each cow was biopsied two or three times during the experiment with five to eight mammary tissue biopsies on d 0, 7, 14, 21, 28, 35, 49, and 130. Mammary tissue preinjection had abundant connective and adipose tissues with limited lobuloalveolar structures. Beginning on d 7, there was decreased stroma, increased epithelial cell area, increased lobuloalveolar architecture, plus the accumulation of intracellular and intraluminal secretions which were high in lipid droplets. From d 7 through 35, these changes were progressive although variable among cows. Changes in activities of enzymes and concentrations of ribonucleic acid and deoxyribonucleic acid were gradual during this time but essentially paralleled histological development. Tissue samples during lactation (d 49 and 130) showed increased histological and biochemical development; development was maximal for d 130 samples. Fourteen of 15 cows that lactated had mean daily yields of milk more than 5 kg and yields of milk of 12 cows with projected or actual 305-d lactations were 63.0% of that during their previous natural lactations. Reasons for less yields of milk and for varied patterns of tissue development were not identified nor explained by concentrations of several selected hormones in plasma.