Technical note: Automatic evaluation of infrared thermal images by computerized active shape modeling of bovine udders challenged with Escherichia coli.

Mastitis causes substantial economic losses and animal suffering in the dairy industry. The trend toward larger herd sizes complicates the monitoring of udder health in individual animals. Infrared thermography has successfully been used for early mastitis detection. However, manual thermogram analysis is time consuming and requires a skilled examiner, and automated image processing has not been tested. The aim of this study was to determine whether automatic evaluation of thermograms showed results comparable to those of manual evaluation of thermograms. Five healthy cows underwent an intramammary challenge with Escherichia coli to induce clinical mastitis. Multiple udder thermograms were taken every 2 h for 24 h before and after the challenge, resulting in 4,143 images in total. All images were evaluated using image recognition software (automatically) and a polygon tool (manually) to calculate the average and maximum surface temperatures. Because of the slightly different regions of interest, temperatures ascertained from the thermograms using the automatic method were consistently lower than those ascertained using the manual method. However, average udder surface temperatures evaluated using both methods were strongly correlated (r = 0.98 in the left hindquarter, and r = 0.99 in the right hindquarter) and showed maximum temperature peaks at the same time, 13 and 15 h after intramammary challenge. In the receiver operating characteristic analysis, both methods provided good results for sensitivity and specificity in detecting clinical E. coli-induced mastitis at different threshold values. For automatically evaluated maximum right hindquarter temperature, sensitivity was 93.75% and specificity was 94.96%, and for manually evaluated maximum right hindquarter temperature, sensitivity was 93.75% and specificity was 96.40%. Thus, automatic thermogram evaluation is a promising tool for automated mastitis detection.

Ankyrin-1 mutations are a major cause of dominant and recessive hereditary spherocytosis.

Hereditary spherocytosis (HS) is the most common inherited haemolytic anaemia in Northern Europeans. The primary molecular defects reside in the red blood cell (RBC) membrane, particularly in proteins that link the membrane skeleton to the overlying lipid bilayer and its integral membrane constituents. Ankyrin-1 is the predominant linker molecule. It attaches spectrin, the major skeletal protein, to the cytoplasmic domain of band 3, the RBC anion exchanger. Two-thirds of patients with HS have combined spectrin and ankyrin-1 deficiency; deficiency of band 3 occurs in about 15 to 20% (ref.1). These data suggest that ankyrin-1 or band 3 defects may be common in HS. To test this we screened all 42 coding exons plus the 5' untranslated/promoter region of ankyrin-1 and the 19 coding exons of band 3 in 46 HS families. Twelve ankyrin-1 mutations and five band 3 mutations were identified. Missense mutations and a mutation in the putative ankyrin-1 promoter were common in recessive HS. In contrast, ankyrin-1 and band 3 frameshift and nonsense null mutations prevailed in dominant HS. Increased accumulation of the normal protein product partially compensated for the ankyrin-1 or band 3 defects in some of these null mutations. Our findings indicate that ankyrin-1 mutations are a major cause of dominant and recessive HS (approximately 35 to 65%), that band 3 mutations are less common (approximately 15 to 25%), and that the severity of HS is modified by factors other than the primary gene defect.

Interactions of spectrin in hereditary elliptocytes containing truncated spectrin beta-chains.

An abnormal spectrin, in which one subunit is truncated, has been detected in a large German family. The inheritance is autosomal dominant. The affected members of the family suffer in widely varying degree from a microcytic hemolytic anemia. The red cell morphology varies correspondingly from smooth elliptocytes to predominantly poikilocytes. The abnormal spectrin makes up approximately 30% of the total and is almost entirely present as the dimer. The truncated chain is not phosphorylated by the endogenous cAMP-independent kinase, and it has been identified as a chain of beta-type, using monoclonal antibodies. Because a univalent terminal spectrin alpha-chain fragment will bind to normal dimers with an association constant lower by only a factor of two than that for the self-association of the dimers, it would be expected that the mutant dimers (alpha beta') would readily enter into an association with normal (alpha beta) dimers to give alpha 2 beta beta' tetramers (though not with each other). In dilute solution this is indeed observed, and the diminution in tetramer concentration when 30% of normal spectrin is replaced by alpha beta' dimers, amounts to only a small proportion. Moreover, in the membrane skeleton, if there is pairwise apposition of dimer units, only 9% of pairings will be between units that cannot associate. We have shown that the failure of alpha beta' dimers to enter into heterologous associations in situ is not due to the elimination of the ankyrin binding site near the truncated end of the beta-chain: this site is fully functional, as judged by rebinding to spectrin-depleted vesicles. When the spectrin is extracted from the membrane in the cold, the material released initially consists almost entirely of alpha beta' dimers; when the spectrin of normal membranes is partly dissociated to dimers in situ by warming at low ionic strength, extraction in the cold then leads similarly to much more rapid release of the dimer than of the tetramer. The similar rates of liberation of normal and abnormal dimer make it unlikely that the interaction of the latter with the membrane is in any way defective. When mixtures of alpha beta and alpha beta' dimers are bound to spectrin-depleted inside-out membrane vesicles from normal cells and tetramers are allowed to form by equilibration at 30 degrees C, the proportion of the abnormal species appearing in the tetramer is much lower than would be expected on a statistical basis. The relation of the self-association equilibrium on the membrane to that of spectrin in dilute solution is analyzed.

Organization of enzymes of glycolysis and of glutathione metabolism in human red cell membranes.

1) The activities of 16 enzymes of glycolysis and of glutathione metabolism were determined in intact human red cell membranes (ghosts) which were prepared by hypotonic hemolysis. 2) Enzymes and hemoglobin of the ghosts were resolved by two toluene extractions. Only the four enzymes hexokinase, fructose-bisphosphate aldolase, glyceraldehyde-phosphate dehydrogenase and pyruvate kinase could not be released completely from the ghosts. 3) The residual membrane fraction, which was obtained after the toluene extraction of ghosts prepared at 30 imOsM, contained 0.02% of the original hemoglobin content of the red cell. Between 6.5 and 23% of the hemolysate activities of glyceraldehyde-phosphate dehydrogenase, phosphoglycerate kinase, pyruvate kinase and fructose-bisphosphate aldolase were detected in this fraction after mechanical disruption. 4) Sonication of intact ghosts increased the activities of fructose-bisphosphate aldolase, pyruvate kinase and phosphoglycerate kinase. 5) In "white" ghosts prepared at 5 imOsM phosphate buffer which contained 0.5% of the original hemoglobin the activities of fructose-bisphosphate aldolase and glyceraldehyde-phosphate dehydrogenase were detected at high levels. The activities of pyruvate kinase and phosphoglycerate kinase were low in these preparations. 6) The results indicate that one part of all enzymes is loosely attached to the inner surface of the membrane as is hemoglobin. A second part, the "cryptic enzyme activity", is available after resolving by toluene. A residual part of four enzymes is firmly bound to the membrane. Two of them (fructose-bisphosphate aldolase and glyceraldehyde-phosphate dehydrogenase) are oriented toward the inner surface of the membrane, whereas pyruvate kinase and phosphoglycerate kinase are hidden in the lipid core of the membrane.

Hyperphagia in children with craniopharyngioma is associated with hyperleptinaemia and a failure in the downregulation of appetite.

Patients with craniopharyngioma frequently suffer from severe obesity. Leptin induces an inhibition of appetite via hypothalamic receptors. This study was undertaken to investigate whether a relationship exists between serum leptin levels and pituitary/hypothalamic lesions in craniopharyngioma patients. Serum leptin levels were evaluated by RIA in 14 patients (age 7-21 years; 7 females, 7 males) after they had undergone neurosurgical treatment for craniopharyngioma. Normal controls had a positive correlation between leptin levels and body mass index (BMI) with higher levels in the females than in the males. Significantly elevated leptin levels with respect to BMI were found in 11 craniopharyngioma patients who had been affected by a suprasellar tumour, whereas 3 patients with an intrasellar tumour had lower, almost normal serum leptin levels. Our data suggest that craniopharyngioma patients develop hypothalamic obesity because their hypothalamic structures are insensitive to endogenous leptin. The elevated serum leptin concentrations found only in patients with a suprasellar tumour may be explained by a disturbed feedback mechanism from the hypothalamic leptin receptors to the adipose tissue.

Red cell pyruvate kinase deficiency: an optimised assay.

As a result of using a variety of assay techniques, rather different values for the rate of the pyruvate kinase (PK) reaction were obtained. Kinetic measurements performed with red cell PK using the stopped-flow as well as the "classical" spectrophotometric method demonstrated that only the initiation of the reaction with ADP yields reproducible results in the standard assay. Moreover, the influence of substrate concentrations, activator concentrations, ionic strength, buffer system, temperature and duration of preincubation were thoroughly investigated and were shown to play an important role. As a consequence of our kinetic measurements, a modified optimised assay composition together with new normal values for erythrocyte PK activity in haemolysate are presented. For better characterisation and comparability of PK variants, we suggest the use of the optimised assay composition described and that the reaction be initiated with ADP.

Analysis of high resolution transmission electron microscope images of crystalline-amorphous interfaces.

For the analysis of images of homogeneous crystalline-amorphous interfaces we propose to average them along the interface obtaining the averaged interface image or the averaged intensity profile. Due to averaging, contrast components with the periodicity of the crystalline area of the image are extracted. Thus, the contrast features originating from the random overlap of the projected potentials of atoms in the amorphous layer are suppressed. It is shown that averaged images can be simulated by the multi-slice method using the novel approach to model the near interfacial amorphous structure by its mean atomic density distribution in front of the crystalline boundary. The crystalline structure is represented by its known atomic positions. We apply the proposed method to the investigation of the near interfacial short-range order in the c-Si/ a-Ge crystalline-amorphous interface.

[Cytologic diagnosis of cancer of the urinary bladder, reliability, pitfalls and grading (author's transl)]. / Treffsicherheit, Grading and Ursachen diagnostischer Fehler bei der zytologischen Diagnose des Blasenkarzinoms.

The cytologic analysis of bladder washings was positive in 59 (67%) of 88 patients with carcinoma of the bladder. About 50% of the false negative cytologic reports were shown to be misinterpretations. The cytologic report was positive in six of 29 cases, in which non-neoplastic lesions, were established by histology. In two cases the positive cytologic report was confirmed by repeat biopsies. In two further cases no neoplasm was histologically verified by repeat biopsy, but the positive cytologic findings were supported by positive results obtained by impulscyto-photometric studies of the nuclear DNA-content in cells of the bladder washings. Two positive cytologic reports were definitely considered as true false positive. Cytologic malignancy grading was attempted without knowledge of the original cytologic and histologic diagnoses. Cytologic and histologic grading were congruent in 68% of cases with positive cytologic report, in 16% of these cases, however, the cytologic grading revealed a higher grade of malignancy. Retrospectively in 27% of the patients examined the cytologic report proved to be clinical value concerning the follow-up as well as the histologic re-examination or the therapy.

[DNA profile, recurrence rate and progression of superficial G2 cancer of the urinary bladder]. / DNA-Profile, Rezidivrate und Progression beim oberflächlichen G2-Karzinom der Harnblase.

In a retrospective study the prognostic value of flow cytometric DNA analysis was studied in 114 newly diagnosed cases with superficial bladder tumours. G1-tumours were generally diploid, G3-tumours almost exclusively aneuploid. G2-tumours exhibited a bimodal distribution with 50% diploid and 50% aneuploid cases respectively. Recurrence and progression correlated systematically with the ploidy level. Furthermore, the degree of aneuploidy is predictive for proneness to progression.

[Importance of impulse cytophotometry in the diagnosis and prognosis of prostate cancer]. / Bedeutung der Impulszytophotometrie für Diagnose und Prognose des Prostatakarzinoms.

In 102 cases with hypertrophy and 73 patients with carcinoma of the prostate, all histologically proven, fine-needle biopsies were done and the obtained material investigated by conventional cytology as well as by flow-through cytophotometric determination of DNA content of the nuclei. In prostatic cancer the DNA determinations showed less diagnostic precision than conventional cytology. However as an additional diagnostic procedure the flow-through cytophotometry was able to improve the results of cytologic diagnostic for 10% in all cases with histologically proven cancer of all grades. In hypertrophy of prostate, the exclusion of carcinoma could be done by DNA determination with higher security than by conventional cytology. Flow-through cytophotometry is therefore considered a useful additional test in primary diagnostic and grading of prostatic cancer. In addition it is a method to evaluate the efficacy of drug therapy.