Nocturnal river water temperatures: Spatial and temporal variations.

Nocturnal water temperature (Tw) affects the behaviour of aquatic biota and metabolism of whole rivers. However, night-time water temperature (nTw) is poorly understood because spot samples are typically taken during daylight hours, or Tw series are aggregated in ways that mask sub-daily properties. This paper examines 15-minute measurements of Tw and air temperature (Ta) collected at 36 sites in the Rivers Dove and Manifold, English Peak District. Data were stratified by day and night then analysed using hysteresis, auto-correlation and logistic regression techniques. Daily hysteresis loops show lagged responses between nTw and previous daylight air temperatures (dTa), plus the influence of groundwater and discharge variations. Logistic regression models were modified using a seasonal factor and explained between 80 and 94% of the variance in daily maximum nTw; minimum nTw were predicted with less skill, particularly for headwater sites in summer. Downstream variations in model parameters also reflect the influence of groundwater and/or riparian shade, and prevailing weather conditions. A case is presented where an intense summer storm resulted in the propagation of a thermal wave that produced maximum Tw at some sites during hours of darkness. Hence, our findings show that Tw management by riparian shade has to be seen in a catchment wide context, with anticipated benefits normalised for weather variability, extreme rainfall events, local influence of groundwater, and channel structures.

Natural history of nonketotic hyperglycinemia in 65 patients.

BACKGROUND: Glycine encephalopathy, also known as nonketotic hyperglycinemia (NKH), is an autosomal recessive disorder caused by a defect in the glycine cleavage system. NKH is classically associated with neonatal apnea, lethargy, hypotonia, and seizures, followed by severe psychomotor retardation in those who survive. METHODS: To determine the natural history of NKH, the authors mailed a 44-question survey to 170 households in the International NKH Family Network. RESULTS: Data for 65 patients (36 boys, 29 girls) were collected from 58 families. One-third of the subjects died; 8 girls died during the neonatal period, and 14 patients died thereafter (2 girls, 12 boys). Median age of death for boys was 2.6 years vs <1 month for girls (p = 0.02). Mean birth weight and length, occipitofrontal circumference, and gestation duration were normal. Two-thirds of infants were ventilated during the neonatal period; of these, 40% died. Ninety percent had confirmed seizures, 75% during the first month of life. Interestingly, three NKH patients never developed seizures. An abnormal corpus callosum and/or hydrocephalus were associated with especially poor gross motor and speech development. Of 25 patients living > or =3 years, 10 were able to walk and say/sign words; all were boys. In six families with more than one affected child, disease course and mortality were similar within each family. CONCLUSIONS: This study reveals a striking and unexpected gender difference in mortality and developmental progress. Of the two-thirds of nonketotic hyperglycinemia patients surviving the newborn period, up to 20% (mostly boys) may learn to walk and communicate by saying or signing words.

Glycine encephalopathy (nonketotic hyperglycinaemia) : review and update.

This article summarizes data and issues covered in the workshop on Glycine Encephalopathy using headings that cover important topics in our present knowledge of this disease.

Three novel deletions in the alanine:glyoxylate aminotransferase gene of three patients with type 1 hyperoxaluria.

We describe three novel deletions in the human AGT gene in three patients with primary hyperoxaluria type 1, an autosomal recessive disease resulting from a deficiency of the liver peroxisomal enzyme, alanine glyoxylate aminotransferase (AGT; EC 2.6.1.44). A deletion of 4 nucleotides in the exon 6/intron 6 splice junction (679-IVS6+2delAAgt) is expected to cause missplicing. It would also code for a K227E missense alteration in any mRNA successfully spliced. A 2-bp deletion in exon 11 (1125-1126del CG, cDNA) results in a frameshift. A deletion of at least 5-6 kb, EX1 EX5del, spanned exons 1-5 and contiguous upstream sequence. All three deletions are heterozygous with previously documented missense mutations; the intron 6 deletion with F152I, the exon 11 deletion with G82E, and EX1 EX5del with the common mistargeting mutation, G170R.

Nonketotic hyperglycinemia (glycine encephalopathy): laboratory diagnosis.

Nonketotic hyperglycinemia (NKH) is an autosomal recessive disorder of glycine metabolism caused by a defect in the glycine cleavage enzyme complex (GCS). GCS is a complex of four proteins encoded on four different chromosomes. In classical neonatal NKH, levels of cerebrospinal fluid (CSF) glycine and CSF/plasma glycine ratio are very high but the CSF results, in particular, may be more difficult to interpret in later-onset, milder, or otherwise atypical NKH. Enzymatic confirmation of NKH requires a liver sample. Delineation of which protein of the complex is defective is necessary to screen for mutations in the appropriate gene. Except for Finnish NKH patients, few recurrent mutations have yet been found, although analysis of the P-protein gene (the site of the defect in the majority of patients) is at an early stage. Prenatal diagnosis by GCS assay in chorionic villus biopsies is not completely reliable and will be replaced by molecular analysis in families where the mutations are known.

Recurrent mutations in P- and T-proteins of the glycine cleavage complex and a novel T-protein mutation (N145I): a strategy for the molecular investigation of patients with nonketotic hyperglycinemia (NKH).

Screening a DNA bank from 50 patients with enzymatic confirmation of their diagnosis of nonketotic hyperglycinemia gave allele frequencies of 5% for R515S of P-protein (glycine decarboxylase) and 7% for R320H of T-protein (aminomethyltransferase). In a previous report we found that 3% of the same patient alleles were positive for T-protein IVS7-1G>A. In total, testing for these three mutations identified 15% of alleles and positive results (one or two mutations) were found in 11 of the 50 patients. In addition, a novel point mutation in T-protein, N145I, was found in a single case and a PCR/restriction enzyme assay was developed for its detection.

Identification of the first reported splice site mutation (IVS7-1G-->A) in the aminomethyltransferase (T-protein) gene (AMT) of the glycine cleavage complex in 3 unrelated families with nonketotic hyperglycinemia.

A novel splice site mutation (IVS7-1G-->A) in the T-protein gene (aminomethyltransferase, or AMT) of the glycine cleavage enzyme complex was found in a patient with nonketotic hyperglycinemia (NKH). A PCR/restriction enzyme method to detect this mutation was used to screen 100 NKH alleles and identified the mutation in three unrelated families.

Biochemical and molecular investigations of patients with nonketotic hyperglycinemia.

The investigation of 14 unrelated patients with nonketotic hyperglycinemia led to the identification of mutations in 4 cases. Patients were initially categorized into probable P- or T-protein defects of the glycine cleavage enzyme complex, by the use of the glycine exchange assay without supplemental H-protein, then screened for mutations in the P-protein and T-protein genes, respectively.

Non-concordance of CVS and liver glycine cleavage enzyme in three families with non-ketotic hyperglycinaemia (NKH) leading to false negative prenatal diagnoses.

We report three false negative prenatal diagnostic results, using direct measurement of glycine cleavage enzyme activity in uncultured chorionic villus tissue from 290 pregnancies at risk for non-ketotic hyperglycinaemia (NKH). Testing was done by two centres: Vancouver, Canada and Lyon, France. One false negative result had activity near the lower limit of the normal range but two samples gave completely normal results well within the control range. All three pregnancies continued and the three children were born affected with NKH. Because of the first result, we now counsel that there is a grey zone of uninterpretable activity where affected and normal enzyme values overlap. Because of the other two results we now counsel that there is an approximately 1% chance of a pregnancy with a normal CVS activity resulting in an affected child. The clinical and biochemical findings in the three families are discussed.

Incidence of inborn errors of metabolism in British Columbia, 1969-1996.

OBJECTIVE: To determine how many children with specific types of inborn errors of metabolism are born each year in British Columbia, Canada. This population provides a relatively unique setting for collection of accurate and uniform incidence data because the diagnoses are all made through one laboratory in a population with universal access to government-funded medical care. METHODOLOGY: We used the records of the Biochemical Diseases Laboratory, Children's Hospital, Vancouver (the central referral point for all metabolic diagnoses in British Columbia) to identify all patients diagnosed with the metabolic diseases defined below. We obtained incidence figures by including only the children diagnosed with the diseases covered in this article who were confirmed as having been born within the province for the years 1969 to 1996. The diseases covered were diseases of amino acids, organic acids, the urea cycle, galactosemia, primary lactic acidoses, glycogen storage diseases, lysosomal storage diseases, and diseases involving specifically peroxisomal and mitochondrial respiratory chain dysfunction. Because the technology needed for diagnosis of specific disease groups was in place at different times our data for the different disease groups correspond to different time frames. We have also adjusted the time frames used to allow for the likelihood that some diseases may not come to medical attention for some time after birth. For instance the incidence of amino acid diseases was assessed throughout the whole of this time frame but the incidence of peroxisomal diseases was restricted to 1984 to 1996 because this was the time frame during which the technology needed for diagnosis was in place and reliable. Most disease group statistics included at least 400 000 births. RESULTS: The overall minimum incidence of the metabolic diseases surveyed in children born in British Columbia is approximately 40 cases per 100 000 live births. This includes phenylketonuria (PKU) and galactosemia which are detected by a newborn screening program. Metabolic diseases, which were not screened for at birth, ie, those with PKU and galactosemia subtracted from the total, have a minimal incidence of approximately 30 cases per 100 000 live births. This diagnostic dilemma group would present to pediatricians for diagnosis. Not all metabolic diseases have been surveyed and our data are restricted to the following metabolic disease groups. Approximately 24 children per 100 000 births (approximately 60% of the total disease groups surveyed) have a disease involving amino acids (including PKU), organic acids, primary lactic acidosis, galactosemia, or a urea cycle disease. These children all have metabolic diseases involving small molecules. Approximately 2.3 children per 100 000 births ( approximately 5%) have some form of glycogen storage disease. Approximately 8 per 100 000 births (20%) have a lysosomal storage disease; approximately 3 per 100 000 births (7%-8%) have a respiratory chain-based, mitochondrial disease and approximately 3 to 4 per 100 000 (7%-8%) of births have a peroxisomal disease. The diseases involving subcellular organelles represent approximately half of the diagnostic dilemma group. The incidence of each of the specific diseases diagnosed, including apparently rare diseases such as nonketotic hyperglycinemia, is to be found in the text. The metabolic diseases reported in this survey represent over 10% of the total number of single gene disorders in our population. CONCLUSIONS: Our data provide a good estimate of metabolic disease incidence, for the disease groups surveyed, in a predominantly Caucasian population. Incidence data for metabolic diseases are hard to collect because in very few centers are diagnoses centralized for a population with uniform access to modern health care and this has been the case for our population during the course of the study. (ABSTRACT TRUNCATED)

Atypical nonketotic hyperglycinemia with normal cerebrospinal fluid to plasma glycine ratio.

The diagnosis of nonketotic hyperglycinemia is considered to depend upon the presence of increased cerebrospinal fluid glycine and an increased cerebrospinal fluid to plasma glycine ratio. We studied two siblings who have the neurologic and peripheral biochemical features of the atypical variant of nonketotic hyperglycinemia but have normal cerebrospinal fluid glycine and cerebrospinal fluid to plasma glycine ratios. The proband had reduced liver glycine cleavage system activity of 17% and 21% of mean normal values, confirmed in two independent laboratories. Her lymphoblast glycine cleavage system activity was normal. Nonketotic hyperglycinemia can be present in the absence of increased cerebrospinal fluid glycine. Measurement of liver glycine cleavage system activity is indicated when nonketotic hyperglycinemia is suggested by clinical features and peripheral glycine levels but cerebrospinal fluid glycine is normal.

Identification of 6 new mutations in the iduronate sulfatase gene. Mutation in brief no. 233. Online.

Mucopolysaccharidosis type II (Hunter syndrome) is an X-linked lysosomal storage disorder caused by a deficiency of the enzyme iduronate-2-sulfatase. We sequenced genomic DNA and RT-PCR products in the iduronate sulfatase (IDS) gene in 6 unrelated patients with Hunter syndrome to assess genotype/phenotype relationships and offer carrier testing where required. Six novel mutations were identified: four missense mutations, one four-base pair deletion (596-599delAACA) and a cryptic splice site mutation. Three of the missense mutations were significant amino acid substitutions (S143F, S491F, E341K) of which the latter two involve amino acids conserved amongst sulfatase enzymes. The patients identified with these mutations all had a severe clinical phenotype. One missense mutation with a minimal amino acid substitution (H342Y), in a non-conserved region of the gene, was associated with a mild clinical phenotype. We identified a novel cryptic splice site (IVS5+934G>A) with some normal (wild type) mRNA processing. We predict that the normal mRNA product confered some residual functional enzyme, resulting in a mild phenotype associated with the absence of overt central nervous system disease.

A protocol for detection of mitochondrial DNA deletions: characterization of a novel deletion.

OBJECTIVES: To develop a protocol capable of identifying deletions in mitochondrial DNA and use it to identify the breakpoints of a mtDNA deletion in a patient with chronic progressive external ophthalmoplegia (CPEO). DESIGN AND METHODS: Deletions in mtDNA were identified by a combination of long range PCR and Southern blotting. The precise breakpoints were determined by automated DNA sequencing. RESULTS: A series of DNA samples from patients with suspected mitochondrial disease was subjected to a protocol, which combines long range PCR and Southern blotting. We found a unique deletion in a patient with CPEO and we identified the precise location of this deletion through DNA sequencing. CONCLUSIONS: Long range PCR has the advantages of speed, minimal samples requirements, and sensitivity. Southern blotting is better able to evaluate heteroplasmy and detect duplications. We suggest a protocol that enables us to identify precisely the breakpoints in a unique mutation of mtDNA in a patient with CPEO.

Human mucopolysaccharidosis IIID: clinical, biochemical, morphological and immunohistochemical characteristics.

Mucopolysaccharidosis IIID (MPS IIID) is one of the rarest of the MPS-III syndromes. To date, the clinical manifestations of 10 patients have been reported, the deficient N-acetylglucosamine 6-sulfatase (G6S) enzyme has been purified, and the G6S gene has been cloned, sequenced and localized. However, morphological manifestations of this condition have not been reported and the pathogenesis of the severe neurological deficits remains an enigma. In this paper we describe and correlate the clinical, biochemical and pathological observations for 2 cases of MPS IIID. We used monoclonal antibodies against heparan sulfate (HS) and GM2-ganglioside, thin layer chromatography, mass spectrometry, and morphological techniques to demonstrate the nature and the distribution of the uncatabolized substrates. The majority of the cells in various tissues showed morphological changes expected with lysosomal storage of HS. The central nervous system (CNS) was most severely affected because of the secondary storage of GM2 and GM3 gangliosides in addition to the primary accumulation of HS. The extent as well as the distribution of the diverse storage materials varied within and among different neurons as observed in MPS-III A, B, and C syndromes. This study supports the hypothesis that the neurological dysfunction and neurodegeneration common to the Sanfilippo syndromes is, in part, due to the secondary metabolic perturbations induced by HS accumulation.

Murine mucopolysaccharidosis type I: targeted disruption of the murine alpha-L-iduronidase gene.

Mucopolysaccharidosis type I (MPS I) is considered to represent the prototypical mucopolysaccharide storage disorder. Although a spectrum of severity is seen within the MPS I subgroup, Hurler syndrome represents the most severe and frequent manifestation of MPS I. We describe here the generation of a murine model for Hurler syndrome by targeted disruption of the murine Idua gene. Homozygous Idua -/- mice have no detectable alpha-L-iduronidase enzyme activity and show increased urinary glycosaminoglycan levels. Although normal appearing at birth, Idua -/- mice develop a flattened facial profile and thickening of the digits discernible by 3 weeks of age. No obvious growth deficiency nor mortality is seen within the first 20 weeks of life. Radiographs reveal anterior flaring of the ribs and thickening of the facial bones as early as 4 weeks of age with more extensive dysostosis detectable by 15 weeks of age. At 4 weeks of age, lysosomal storage is noted primarily within reticuloendothelial cells with abundant lysosomes noted in Kupffer cells, splenic sinusoidal lining cells, and glial cells. More widespread lysosomal storage is noted by 8 weeks of age in hepatocytes, chondrocytes, neurons, as well as renal tubular cells. Thus, targeted disruption of the murine Idua locus has produced a murine strain representative of the severe form of MPS I. This model should permit detailed evaluation of the pathophysiology of lysosomal storage disorders and provide a small animal model for the testing and development of enzyme replacement and gene therapy regimes.

The ultrastructure of skin from a patient with mucopolysaccharidosis IIID.

Mucopolysaccharidosis IIID (MPS-IIID) is the rarest of the MPS-III syndromes. It is caused by deficient activity of lysosomal N-acetylglucosamine-6-sulfatase (G6S). To date, the clinical and biochemical features of seven patients with MPS-IIID have been reported, but no biopsy or autopsy findings have been described. The purpose of this report is to define the ultrastructure of affected cells seen in a skin biopsy from a 14-year-old boy. The child presented with progressive mental deterioration, hyperactivity and mild to moderate dysmorphism. The diagnosis of a mucopolysaccharidosis was suggested, but the initial urine analyses were negative for elevated mucopolysaccharides, and only the third analysis showed abnormal excretion of heparan sulfate. Because of the diagnostic difficulties posed by this case, a skin biopsy was performed for morphological and biochemical studies. Numerous vacuoles were noted in Schwann cells, fibroblasts, smooth muscle cells, eccrine gland and ductal epithelium in resin-embedded sections stained with toluidine blue. Ultrastructurally, many lysosomes were distended with abundant, fibrillar material. Occasionally, lamellated membranous structures were present within the same lysosomes. These findings are consistent with those seen in other forms of MPS, in which the lysosomal storage occurs predominantly, but not exclusively, in mesenchymal cells. Furthermore, deficient activity of G6S was confirmed in cultured skin fibroblasts. This study demonstrates that electron microscopy of skin biopsies is a useful method for identification of patients with clinical features of MPS-IIID whether or not heparan sulfaturia is present.

DNA-based diagnosis of arylsulfatase A deficiencies as a supplement to enzyme assay: a case in point.

OBJECTIVE: To identify the molecular basis of arylsulfatase A deficiency in a family at risk for metachromatic leukodystrophy (MLD) and determine the genetic risk in the offspring. METHODS: Mutations in the arylsulfatase A gene were identified by PCR amplification and restriction enzyme digestion. Individuals had previously been tested for arylsulfatase A activity. RESULTS: Assays of arylsulfatase A activity had resulted in ambiguous results for MLD carrier identification. DNA analysis clearly identified two MLD mutations in the family, and an unsuspected arylsulfatase A pseudodeficiency. The DNA information immediately clarified the MLD risk for the family and confirmed that a newborn with low arylsulfatase A activity was unaffected. CONCLUSIONS: The overlap between activities for various combinations of MLD and pseudodeficiency alleles and the variability inherent in the assay of arylsulfatase A complicate the interpretation of activity levels in families at risk for MLD. Use of simple molecular biological tests for pseudodeficiency and the common MLD mutations in combination with the enzyme data can facilitate carrier identification and prenatal diagnosis.

Nonketotic hyperglycinemia: atypical clinical and biochemical manifestations.

A 16-year-old boy had intermittent chorea, delirium, and vertical gaze palsy precipitated by febrile illness. Nonketotic hyperglycinemia was confirmed by measurement of liver and lymphoblast glycine cleavage enzyme activity. Deficient but residual enzyme activity was demonstrated in both tissues, possibly accounting for the mild phenotype. Confirmation of an atypical variant of nonketotic hyperglycinemia with residual glycine cleavage enzyme activity has important implications for diagnosis and treatment.