Low Mannan-binding lectin serum levels are associated with complicated Crohn's disease and reactivity to oligomannan (ASCA)

OBJECTIVES: Mannan-binding lectin (MBL) acts as a pattern-recognition molecule directed against oligomannan, which is part of the cell wall of yeasts and various bacteria. We have previously shown an association between MBL deficiency and anti-Saccharomyces cerevisiae mannan antibody (ASCA) positivity. This study aims at evaluating whether MBL deficiency is associated with distinct Crohn's disease (CD) phenotypes. METHODS: Serum concentrations of MBL and ASCA were measured using ELISA (enzyme-linked immunosorbent assay) in 427 patients with CD, 70 with ulcerative colitis, and 76 healthy controls. CD phenotypes were grouped according to the Montreal Classification as follows: non-stricturing, non-penetrating (B1, n=182), stricturing (B2, n=113), penetrating (B3, n=67), and perianal disease (p, n=65). MBL was classified as deficient (<100 ng/ml), low (100-500 ng/ml), and normal (500 ng/ml). RESULTS: Mean MBL was lower in B2 and B3 CD patients (1,503+/-1,358 ng/ml) compared with that in B1 phenotypes (1,909+/-1,392 ng/ml, P=0.013). B2 and B3 patients more frequently had low or deficient MBL and ASCA positivity compared with B1 patients (P=0.004 and P<0.001). Mean MBL was lower in ASCA-positive CD patients (1,562+/-1,319 ng/ml) compared with that in ASCA-negative CD patients (1,871+/-1,320 ng/ml, P=0.038). In multivariate logistic regression modeling, low or deficient MBL was associated significantly with B1 (negative association), complicated disease (B2+B3), and ASCA. MBL levels did not correlate with disease duration. CONCLUSIONS: Low or deficient MBL serum levels are significantly associated with complicated (stricturing and penetrating) CD phenotypes but are negatively associated with the non-stricturing, non-penetrating group. Furthermore, CD patients with low or deficient MBL are significantly more often ASCA positive, possibly reflecting delayed clearance of oligomannan-containing microorganisms by the innate immune system in the absence of MBL.

A mutation in the human ortholog of the Saccharomyces cerevisiae ALG6 gene causes carbohydrate-deficient glycoprotein syndrome type-Ic

Carbohydrate-deficient glycoprotein syndrome (CDGS) represents a class of genetic diseases characterized by abnormal N-linked glycosylation. CDGS patients show a large number of glycoprotein abnormalities resulting in dysmorphy, encephalopathy, and other organ disorders. The majority of CDGSs described to date are related to an impaired biosynthesis of dolichyl pyrophosphate-linked Glc3Man9GlcNAc2 in the endoplasmic reticulum. Recently, we identified in four related patients a novel type of CDGS characterized by an accumulation of dolichyl pyrophosphate-linked Man9GlcNAc2. Elaborating on the analogy of this finding with the phenotype of alg5 and alg6 Saccharomyces cerevisiae strains, we have cloned and analyzed the human orthologs to the ALG5 dolichyl phosphate glucosyltransferase and ALG6 dolichyl pyrophosphate Man9GlcNAc2 alpha1,3-glucosyltransferase in four novel CDGS patients. Although ALG5 was not altered in the patients, a C-->T transition was detected in ALG6 cDNA of all four CDGS patients. The mutation cosegregated with the disease in a Mendelian recessive manner. Expression of the human ALG5 and ALG6 cDNA could partially complement the respective S. cerevisiae alg5 and alg6 deficiency. By contrast, the mutant ALG6 cDNA of CDGS patients failed to revert the hypoglycosylation observed in alg6 yeasts, thereby proving a functional relationship between the alanine to valine substitution introduced by the C-->T transition and the CDGS phenotype. The mutation in the ALG6 alpha1,3-glucosyltransferase gene defines an additional type of CDGS, which we propose to refer to as CDGS type-Ic.

Evaluation of PCR electrospray-ionization mass spectrometry for rapid molecular diagnosis of bovine mastitis

Bovine mastitis, an inflammatory disease of the mammary gland, is one of the most costly diseases affecting the dairy industry. The treatment and prevention of this disease is linked heavily to the use of antibiotics in agriculture and early detection of the primary pathogen is essential to control the disease. Milk samples (n=67) from cows suffering from mastitis were analyzed for the presence of pathogens using PCR electrospray-ionization mass spectrometry (PCR/ESI-MS) and were compared with standard culture diagnostic methods. Concurrent identification of the primary mastitis pathogens was obtained for 64% of the tested milk samples, whereas divergent results were obtained for 27% of the samples. The PCR/ESI-MS failed to identify some of the primary pathogens in 18% of the samples, but identified other pathogens as well as microorganisms in samples that were negative by culture. The PCR/ESI-MS identified bacteria to the species level as well as yeasts and molds in samples that contained a mixed bacterial culture (9%). The sensitivity of the PCR/ESI-MS for the most common pathogens ranged from 57.1 to 100% and the specificity ranged from 69.8 to 100% using culture as gold standard. The PCR/ESI-MS also revealed the presence of the methicillin-resistant gene mecA in 16.2% of the milk samples, which correlated with the simultaneous detection of staphylococci including Staphylococcus aureus. We demonstrated that PCR/ESI-MS, a more rapid diagnostic platform compared with bacterial culture, has the significant potential to serve as an important screening method in the diagnosis of bovine clinical mastitis and has the capacity to be used in infection control programs for both subclinical and clinical disease.

Pathology of sarcoptic mange in red foxes (Vulpes vulpes): macroscopic and histologic characterization of three disease stages

Sarcoptic mange is a highly contagious skin disease that can have a devastating impact on affected wild mammal populations. There are notable variations in the clinical and pathologic picture of sarcoptic mange among species and among conspecifics. However, the origin of these variations is unclear. We propose a classification scheme for skin lesions associated with Sarcoptes scabiei infestation to provide a basis for a subsequent risk factor analysis. We conducted a case-control study focused on macroscopic and histologic examination of the skin, using 279 red foxes (Vulpes vulpes) found dead or shot in Switzerland between November 2004 and February 2006. All animals were submitted to gross necropsy following a detailed protocol. Selection criteria for cases (n=147) vs. controls (n=111) were the presence or absence of mange-like lesions, mite detection by isolation or histologic examination, and serologic testing for S. scabiei antibodies. Characteristic features of mange lesions were scored macroscopically in all foxes and histologically in 67 cases and 15 controls. We classified skin lesions and associated necropsy findings into three types of mange: A) early stage (n=45): focal-extensive skin lesions, thin crusts, mild to moderate alopecia, few mites, numerous eosinophils, and mild lymph node enlargement; B) hyperkeratotic, fatal form (n=86): generalized skin lesions, thick crusts with or without alopecia, foul odor, abundance of mites, numerous bacteria and yeasts, numerous lymphocytes and mast cells, severe lymph node enlargement, and emaciation; C) alopecic, healing form (n=16): focal lesions, no crusts, severe alopecia, hyperpigmentation and lichenification, absence of mites, mixed cell infiltration, and rare mild lymph node enlargement. We hypothesize that after stage A, the animal either enters stage B and dies, or stage C and survives, depending on largely unknown extrinsic or intrinsic factors affecting the host ability to control mite infestation.

Transcript levels of AtMRPs after cadmium treatment: induction of AtMRP3

In yeasts, the ABC-type transporters are involved in vacuolar sequestration of cadmium. In plants, transport experiments with isolated vacuoles indicate that this is also true. In order to know more about the response of AtMRPs, a subclass of Arabidopsis ABC transporters, to cadmium, their expression pattern was analysed using the microchip technology and semi-quantitative reverse transcriptase-polymerase chain reaction. From 15 putative sequences coding for AtMRPs, transcript levels were detected for 14. All were expressed in the roots as well as in the shoots, although at a different level. In 4-week-old Arabidopsis, transcript levels of four AtMRPs were up-regulated after cadmium treatment. In all cases up-regulation was exclusively observed in the roots. The increase of transcript levels was most pronounced for AtMRP3. A more detailed analysis revealed that induction of AtMRP3 could also be observed in the shoot when leaves were cut and cadmium allowed to be taken up in the shoot. In young plantlets, a far higher portion of Cd2+ was translocated in the aerial part compared with adult plants. Consequently, AtMRP3 transcript levels increased in both root and shoot of young plants. This suggests that 7-day-old seedlings do not exhibit such a strict root–shoot barrier as 4-week-old plants. Expression analysis with mutant plants for glutathione and phytochelatin synthesis as well as with compounds producing oxidative stress indicate that induction of AtMRP3 is likely due to the heavy metal itself.

Microbiota at teeth and implants in partially edentulous patients. A 10-year retrospective study.

OBJECTIVE To determine the microbiota at implants and adjacent teeth 10 years after placement of implants with a sandblasted and acid-etched surface. MATERIAL AND METHODS Plaque samples obtained from the deepest sites of 504 implants and of 493 adjacent teeth were analyzed for certain bacterial species associated with periodontitis, for staphylococci, for aerobic gram-negative rods, and for yeasts using nucleic acid-based methods. RESULTS Species known to be associated with periodontitis were detectable at 6.2-78.4% of the implants. Significantly higher counts at implants in comparison with teeth were assessed for Tannerella forsythia, Parvimonas micra, Fusobacterium nucleatum/necrophorum, and Campylobacter rectus. Higher counts of periodontopathogenic species were detectable at implants of current smokers than at those of non-smokers. In addition, those species were found in higher quantities at implants of subjects with periodontitis. The prevalence of Prevotella intermedia, Treponema denticola, C. rectus, and moreover of Staphylococcus warneri might be associated with peri-implant inflammation. Selected staphylococcal species (not Staphylococcus aureus), aerobic gram-negative rods, and yeasts were frequently detected, but with the exception of S. warneri, they did not show any association with periodontal or peri-implant diseases. CONCLUSIONS Smoking and periodontal disease are risk factors for colonization of periodontopathic bacteria at implants. Those bacterial species may play a potential role in peri-implant inflammation. The role of S. warneri needs further validation.

Drought stress and carbon assimilation in a warming climate: Reversible and irreversible impacts

Abstract Global change is characterized by increased {CO2} concentration in the atmosphere, increasing average temperature and more frequent extreme events including drought periods, heat waves and flooding. Especially the impacts of drought and of elevated temperature on carbon assimilation are considered in this review. Effects of extreme events on the subcellular level as well as on the whole plant level may be reversible, partially reversible or irreversible. The photosynthetically active biomass depends on the number and the size of mature leaves and the photosynthetic activity in this biomass during stress and subsequent recovery phases. The total area of active leaves is determined by leaf expansion and senescence, while net photosynthesis per leaf area is primarily influenced by stomatal opening (stomatal conductance), mesophyll conductance, activity of the photosynthetic apparatus (light absorption and electron transport, activity of the Calvin cycle) and {CO2} release by decarboxylation reactions (photorespiration, dark respiration). Water status, stomatal opening and leaf temperature represent a "magic triangle" of three strongly interacting parameters. The response of stomata to altered environmental conditions is important for stomatal limitations. Rubisco protein is quite thermotolerant, but the enzyme becomes at elevated temperature more rapidly inactivated (decarbamylation, reversible effect) and must be reactivated by Rubisco activase (carbamylation of a lysine residue). Rubisco activase is present under two forms (encoded by separate genes or products of alternative splicing of the pre-mRNA from one gene) and is very thermosensitive. Rubisco activase was identified as a key protein for photosynthesis at elevated temperature (non-stomatal limitation). During a moderate heat stress Rubisco activase is reversibly inactivated, but during a more severe stress (higher temperature and/or longer exposure) the protein is irreversibly inactivated, insolubilized and finally degraded. On the level of the leaf, this loss of photosynthetic activity may still be reversible when new Rubisco activase is produced by protein synthesis. Rubisco activase as well as enzymes involved in the detoxification of reactive oxygen species or in osmoregulation are considered as important targets for breeding crop plants which are still productive under drought and/or at elevated leaf temperature in a changing climate.

Molecular mechanisms involved in the regulation of cell polarity and genome stability by the p21 -activated kinase, Shk1, in the fission yeast, Schizosaccharomyces pombe

The essential p21-activated kinase (PAK), Shk1, is a critical component of a Ras/Cdc42/PAK complex required for cell viability, normal cell polarity, proper regulation of cytoskeletal dynamics, and sexual differentiation in the fission yeast, Schizosaccharomyces pombe. While cellular functions of PAKs have been described in eukaryotes from yeasts to mammals, the molecular mechanisms of PAK regulation and function are poorly understood. This study has characterized a novel Shk1 inhibitor, Skb15, and, in addition, identified the cell polarity regulator, Tea1, as a potential biological substrate of Shk1 in S. pombe. Skb15 is a highly conserved WD repeat protein that was discovered from a two-hybrid screen for proteins that interact with the catalytic domain of Shk1. Molecular data indicate that Skb15 negatively regulates Shk1 kinase activity in S. pombe cells. A null mutation in the skb15 gene is lethal and results in deregulation of actin polymerization and localization, microtubule biogenesis, and the cytokinetic machinery, as well as a substantial uncoupling of these processes from the cell cycle. Loss of Skb15 function is suppressed by partial loss of Shk1, demonstrating that negative regulation of Shk1 by Skb15 is required for proper execution of cytoskeletal remodeling and cytokinetic functions. A mouse homolog of Skb15 can substitute for its counterpart in fission yeast, demonstrating that Skb15 protein function has been substantially conserved through evolution. ^ Our laboratory has recently demonstrated that Shk1, in addition to regulating actin cytoskeletal organization, is required for proper regulation of microtubule dynamics in S. pombe cells. The Shk1 protein localizes to interphase and mitotic microtubules, the septum-forming region, and cell ends. This pattern of localization overlaps with that of the cell polarity regulator, Tea1, in S. pombe cells. The tea1 gene was identified by Paul Nurse's laboratory from a screen for genes involved in the control of cell morphogenesis in S. pombe. In contrast to wild type S. pombe cells, which are rod shaped, tea1 null cells are often bent and/or branched in shape. The Tea1 protein localizes to the cell ends, like Shk1, and the growing tips of interphase microtubules. Thus, experiments were performed to investigate whether Tea1 interacts with Shk1. The tea1 null mutation strongly suppresses the loss of function of Skb15, an essential inhibitor of Shk1 function. All defects associated with the skb15 mutation, including defects in F-actin organization, septation, spindle elongation, and chromosome segregation, are suppressed by tea1Δ, suggesting that Tea1 may function in these diverse processes. Consistent with a role for Tea1 in cytokinesis, tea1Δ cells have a modest cell separation defect that is greatly exacerbated by a shk1 mutation and, like Shk1, Tea1 localizes to the septation site. Molecular analyses showed that Tea1 phosphorylation is significantly dependent on Shk1 function in vivo and that bacterially expressed Tea1 protein is directly phosphorylated by recombinant Shk1 kinase in vitro. Taken together, these results identify Tea1 as a potential biological substrate of Shk1 in S. pombe. ^ In summary, this study provides new insights into a conserved regulatory mechanism for PAKs, and also begins to uncover the molecular mechanisms by which the Ras/Cdc42/PAK complex regulates the microtubule and actin cytoskeletons and cell growth polarization in fission yeast. ^

Proceedings of the Nineteenth Annual Biochemical Engineering Symposium

The nineteenth symposium was held at the University of Missouri–Columbia on April 22, 1989. A total of eighteen papers were scheduled for presentation, of which nine were in poster session. Finally, fifteen papers were presented and sixteen were submitted for this proceedings. It was attended by 53 participants from five institutions. A sixth group (from Colorado State University) was kept from attending the symposium due to mechanical problems on the road and we missed them. Since they worked hard at their presentations, I requested CSU-group to submit their papers for the proceedings and I am happy that they did. ContentsMathematical modelling of a flour milling system. K. Takahashi, Y. Chen, J. Hosokoschi, and L. T. Fan. Kansas State University A novel solution to the problem of plasmid segregation in continuous bacterial fermentations. K.L. Henry, R. H. Davis, and A. L. Taylor. University of Colorado Modelling of embryonic growth in avian and reptile Eggs. C.L. Krause, R. C. Seagrave, and R. A. Ackerman. Iowa State University Mathematical modeling of in situ biodegradation processes. J.C. Wu, L. T. Fan, and L. E. Erickson. Kansas State University Effect of molecular changes on starch viscosity. C.H. Rosane and V. G. Murphy. Colorado State University Analysis of two stage recombinant bacterial fermentations using a structured kinetic model. F. Miao and D. S. Kampala. University of Colorado Lactic acid fermentation from enzyme-thinned starch by Lactobacillus amylovorus. P.S. Cheng, E. L. Iannotti, R. K. Bajpai, R. Mueller, and s. Yaeger. University of Missouri–Columbia Solubilization of preoxidized Texas lignite by cell-free broths of Penicillium strains. R. Moolick, M. N. Karim, J. C. Linden, and B. L. Burback. Colorado State University Separation of proteins from polyelectrolytes by ultrafiltration. A.G. Bazzano and C. E. Glatz. Iowa State University Growth estimation and modelling of Rhizopus oligosporus in solid state fermentations. D.-H. Ryoo, V. G. Murphy, M. N. Karim, and R. P. Tengerdy. Colorado State University Simulation of ethanol fermentations from sugars in cheese whey. C.J. Wang and R. K. Bajpai. University of Missouri–Columbia Studies on protoplast fusion of B. licheniformis. B. Shi, Kansas State University Cell separations of non-dividing and dividing yeasts using an inclined settler. C.-Y. Lee, R. H. Davis, and R. A. Sclafani. University of Colorado Effect of·serum upon local hydrodynamics within an airlift column. G.T. Jones, L. E. Erickson, and L. A. Glasgow. Kansas State University Optimization of heterologous protein secretion in continuous culture. A. Chatterjee, W. F. Remirez, and R. H. Davis. University of Colorado An improved model for lactic acid fermentation. P. Yeh, R. K. Bajpai, and E. L. Iannotti. University of Missouri–Columbia

Las yemas, principal reservorio de levaduras entre los órganos aéreos vegetativos y reproductivos de la vid (Vitis vinifera var. Malbec).

El objetivo de esta investigación fue suministrar nueva evidencia acerca del modelo de permanencia de las levaduras en el ciclo natural de la vid. Se efectuó la observación, la medición del número de levaduras y la descripción morfológica de los diferentes órganos aéreos de la vid. Se procedió a la recolección aséptica de muestras a campo, en yema en actividad, yema en reposo, hoja joven, hoja adulta, ritidomis, zarcillo, capullo floral, flor y fruto. Los resultados revelaron dos momentos de máxima población de levaduras: en yema cerrada a fines de otoño y en yema terminal abierta a mediados de verano. La evolución de las levaduras en función de la superficie del fruto mostró poca relación entre ambas variables, por lo que el valor a considerar sería la cantidad de levaduras por baya como unidad. La ritidomis exhibió valores muy uniformes a lo largo del ciclo vegetativo, asumiendo desde esta perspectiva el papel de reservorio de moderada importancia.