Identification of lanthionine synthase C-like protein-1 as a prominent glutathione binding protein expressed in the mammalian central nervous system

Proteomic experiments were performed to identify novel glutathione (GSH) binding proteins expressed in the mammalian central nervous system. Bovine brain lysate was affinity purified using an immobilized glutathione-Sepharose column. Proteins that bound the immobilized glutathione were eluted with free glutathione and identified by one- and two-dimensional electrophoresis coupled with mass spectrometric analysis of tryptic fragments. Major proteins purified by this technique were glutathione S-transferase-mu (GST-mu) and GST-pi and lanthionine synthase C-like protein-1 (LanCL1). LanCL1 is a mammalian homologue of a prokaryotic enzyme responsible for the synthesis of thioether (lanthionine) cross-links within nascent polypeptide chains, yielding macrocyclic proteins with potent microbicidal activity. An antibody against LanCL1 was generated and applied to immunochemical studies of spinal cord tissue from SOD1G93A transgenic mice, a model for amyotrophic lateral sclerosis (ALS), wherein LanCL1 expression was found to be increased at presymptomatic stages of the disease. These results indicate LanCL1 is a glutathione binding protein possibly significant to neurodegenerative disease.     

High-resolution maps of the murine Chromosome 2 region containing the cholesterol gallstone locus, Lith1

The Lith1 region on Chromosome (Chr) 2 contains a gene that markedly affects the prevalence of cholesterol gallstones in inbred mice. We report the high-resolution genetic and radiation hybrid maps of the chromosomal region surrounding Lith1, using three resources: a DNA panel from 188 progeny from two reciprocal backcrosses between C57BL/6 and Mus spretus inbred strains; 423 progeny of an N4 generation from backcrossing the susceptible C57L/J alleles at Lith1 into the resistant AKR/J strain; and the newly developed hamster–mouse T31 radiation hybrid panel. We mapped 17 microsatellite markers in the D2Mit182 to D2Mit14 region and two candidate genes for Lith1, the canalicular bile salt export pump (Bsep) also known as sister of P-glycoprotein (Spgp) and the low-density-lipoprotein-receptor-related gene megalin (Gp330). Both genetic maps were in agreement and ordered the microsatellite markers into a 10.4 ± 1.5 cM region. The high-resolution physical map revealed ordering of microsatellite markers and relative distances between markers in almost complete agreement with the genetic maps. Mapping of Bsep revealed its location on Chr 2, homologous to the human chromosomal position (Nature Genet 20, 233–238, 1998). The radiation hybrid results also provided the highest resolution of the area containing the two candidate genes, which both mapped in the Lith1 region with close linkage, being separated by a distance of only 15 cR₃₀₀₀. The total radiation hybrid map length of the region between D2Mit182 and D2Mit14 was 326 cR₃₀₀₀, suggesting that 31 cR₃₀₀₀ is equivalent to 1 cM in this region of Chr 2. Received: 29 April 1999 / Accepted: 21 July 1999     

Mutation rate dynamics reflect ecological change in an emerging zoonotic pathogen

Mutation rates vary both within and between bacterial species, and understanding what drives this variation is essential for understanding the evolutionary dynamics of bacterial populations. In this study, we investigate two factors that are predicted to influence the mutation rate: ecology and genome size. We conducted mutation accumulation experiments on eight strains of the emerging zoonotic pathogen Streptococcus suis. Natural variation within this species allows us to compare tonsil carriage and invasive disease isolates, from both more and less pathogenic populations, with a wide range of genome sizes. We find that invasive disease isolates have repeatedly evolved mutation rates that are higher than those of closely related carriage isolates, regardless of variation in genome size. Independent of this variation in overall rate, we also observe a stronger bias towards G/C to A/T mutations in isolates from more pathogenic populations, whose genomes tend to be smaller and more AT-rich. Our results suggest that ecology is a stronger correlate of mutation rate than genome size over these timescales, and that transitions to invasive disease are consistently accompanied by rapid increases in mutation rate. These results shed light on the impact that ecology can have on the adaptive potential of bacterial pathogens.     

Technical note: Regression analysis in adult age estimation

Accurate estimation of human adult age has always been a problem for anthropologists, archaeologists and forensic scientists. The main factor contributing to the difficulties is the high variability of physiological age indicators. However, confounding this variability in many age estimation applications is a systematic tendency for age estimates, regardless of physiological indicator employed, to assign ages which are too high for young individuals, and too low for older individuals. This paper shows that at least part of this error is the inevitable consequence of the statistical procedures used to extract an estimate of age from age indicators, and that the magnitude of the error is inversely related to how well an age indicator is correlated with age. The use of classical calibration over inverse calibration is recommended for age estimation. Am J Phys Anthropol 104:259–265, 1997. © 1997 Wiley-Liss, Inc.     

Mitigation of reactive metabolite formation for a series of 3-amino-2-pyridone inhibitors of Bruton’s tyrosine kinase (BTK)

Reactive metabolites have been putatively linked to many adverse drug reactions including idiosyncratic toxicities for a number of drugs with black box warnings or withdrawn from the market. Therefore, it is desirable to minimize the risk of reactive metabolite formation for lead molecules in optimization, in particular for non-life threatening chronic disease, to maximize benefit to risk ratio. This article describes our effort in addressing reactive metabolite issues for a series of 3-amino-2-pyridone inhibitors of BTK, e.g. compound 1 has a value of 459 pmol/mg protein in the microsomal covalent binding assay. Parallel approaches were taken to successfully resolve the issues: establishment of a predictive screening assay with correlation association of covalent binding assay, identification of the origin of reactive metabolite formation using MS/MS analysis of HLM as well as isolation and characterization of GSH adducts. This ultimately led to the discovery of compound 7 (RN941) with significantly reduced covalent binding of 26 pmol/mg protein.     

Fragment size affects plant herbivory via predator loss

Fragmentation and resultant changes in patch size are predicted to alter species diversity and community composition, yet the consequences of these differences for species interactions are poorly understood. Theory predicts that predators are more sensitive to fragmentation than their prey, resulting in greater predator loss in small patches. Predator loss, in turn, is predicted to 1) increase herbivory rates overall, and 2) cause herbivores to shift feeding from plants that act as refugia to those that are preferred forage. We tested these predictions in an old‐field community using two experiments. The first was a large‐scale experiment that included hundreds of arthropod species in fragments of various sizes, and used goldenrod and switchgrass to assess herbivory. Our second experiment manipulated densities of a focal predator species and a focal prey species to determine if changes in densities, rather than other characteristics of fragments, were sufficient to cause the trends observed in the first experiment. We found that predator densities declined in small fragments whereas herbivore densities showed the opposite trend. Total herbivory mirrored herbivore densities by increasing in small patches, and this mean increase was driven by large increases in goldenrod herbivory but declines in switchgrass herbivory. Experimental manipulation of densities confirmed that herbivores preferentially feed on goldenrod, and that predators depress herbivory on goldenrod but have a negligible effect on switchgrass. Our results suggest that fragmentation alters trophic interactions by causing declines in predator densities and increases in herbivore densities, but that feeding preferences of herbivores may generate unequal impacts among plant species.     

Goats in the city: prevalence of <Emphasis Type="Italic">Giardia duodenalis</Emphasis> and <Emphasis Type="Italic">Cryptosporidium</Emphasis> spp. in extensively reared goats in northern India

Background

Various characteristics of goats mean they are highly suitable livestock for backyard rearing by people with limited resources. They are a popular livestock choice in India, where they are often kept to supplement an already scarce income. In these settings, hygiene and sanitation standards tend to be low, and weakens the interface between humans and animals, thus reducing the barrier between them and thereby increasing the likelihood that zoonotic and anthroponotic infections will occur.

Results

This study reports an investigation of the occurrence of Cryptosporidium spp. and Giardia duodenalis in goats being reared in different settings in urban and peri-urban areas in northern India, and addressed the zoonotic potential of these important protozoan parasites shed from goats living close to humans. The overall prevalence of G. duodenalis was 33.8 and 0.5% for Cryptosporidium spp.; the relatively low prevalence of cryptosporidiosis may reflect that most samples were derived from adult animals. The prevalence of G. duodenalis excretion was found to be similar to that reported in other studies. However, although other studies have reported a predominance of non-zoonotic Assemblage E in goats, in this study potentially zoonotic Assemblages predominated [Assemblage A (36%) and Assemblage B (32%)].

Conclusions

The results of this study indicate that in this area where goats and humans are living in close proximity, there may be sharing of intestinal parasites, which can be detrimental for both host species.

     

Ocular herpes: the pathophysiology,management and treatment of herpetic eye diseases

Herpesviruses are a prominent cause of human viral disease, second only to the cold and influenza viruses. Most herpesvirus infections are mild or asymptomatic. However, when the virus invades the eye, a number of pathologies can develop and its associated sequelae have become a considerable source of ocular morbidity. The most common culprits of herpetic eye disease are the herpes simplex virus (HSV), varicella zoster virus (VZV), and cytomegalovirus (CMV). While primary infection can produce ocular disease, the most destructive manifestations tend to arise from recurrent infection. These recurrent infections can wreck devastating effects and lead to irreversible vision loss accompanied by a decreased quality of life, increased healthcare usage, and significant cost burden. Unfortunately, no method currently exists to eradicate herpesviruses from the body after infection. Treatment and management of herpes-related eye conditions continue to revolve around antiviral drugs, although corticosteroids, interferons, and other newer therapies may also be appropriate depending on the disease presentation. Ultimately, the advent of effective vaccines will be crucial to preventing herpesvirus diseases altogether and cutting the incidence of ocular complications.