Electropherotypes and subgroups of group A rotaviruses circulating among diarrhoeic children in Kano, Nigeria.

BACKGROUND: It is estimated that about 600,000 children die annually as a result of severe dehydrating diarrhea caused by rotaviruses. The virus is a double stranded RNA (dsRNA) virus with 11 segments. Group A rotaviruses show a characteristic 4-2-3-2 pattern following electrophoresis. The VP6 subgroups, I and II exist. This work was carried out to study the prevalence of rotavirus infection among children 0-5 years with diarrhea in Kano, and to determine the circulating subgroups and electropherotypes and of the rotavirus isolates. METHODS: Two hundred and eighteen stool specimens from children 0-60 months (198 diarrheic and 20 non-diarrheic) were collected from different hospitals and health care centers in Kano and subjected to group A rotavirus enzyme linked immunosorbent assay (ELISA) to determine presence of group A rotavirus, subgroup ELISA to determine the VP6 subgroups and polyacrylamide gel electrophoresis (PAGE) to determine the electropherotypes present. RESULTS: The long electropherotypes (47.05%) of four variations dominated over the short electropherotype (17.64%). About 11.76% of the isolates were of mixed infection. Dominance of subgroup II (45%) over subgroup I (25%), and the presence of both subgroups I and II (10%) and neither subgroup I nor II (15%) was observed in this study. CONCLUSION: Information on the genomic diversity of the RNA electropherotypes in this region, Kano, is reported in this study.

Activity and tissue-specific expression of lipases and tumor-necrosis factor alpha in lean and obese cats.

Post-heparin plasma activity of lipoprotein lipase (LPL) and hepatic lipase (HL), and fat and muscle activity of LPL were measured in neutered lean and obese cats. Lipoprotein lipase, hormone-sensitive lipase (HSL), and tumor necrosis factor a (TNF) mRNA were measured in muscle and fat tissue with real-time PCR using primers for feline LPL, HSL, and TNF. Lipoprotein lipase plasma and fat activity and fat mRNA levels were significantly lower (50, 80, and 50%, respectively) in obese cats than lean cats, whereas the muscle/fat ratio of LPL was significantly higher in obese compared to lean cats. The activity of HL was not different between the groups. Hormone-sensitive lipase mRNA levels were significantly higher in obese than lean cats. The level of fat TNF also was significantly higher in obese cats than in lean cats, whereas the level in muscle was not different. The lower LPL activity and mRNA expression in fat and the higher LPL and HSL mRNA expression in muscle in obese cats compared to lean cats expectedly favor a redistribution of fatty acids from fat to muscle tissue where they can be deposited or used for energy in times of need. Tumor necrosis factor alpha may regulate this repartitioning process through suppression of adipocyte LPL.

Sir3-dependent assembly of supramolecular chromatin structures in vitro.

Baculovirus-expressed recombinant Sir3p (rSir3p) has been purified to near homogeneity, and its binding to naked DNA, mononucleosomes, and nucleosomal arrays has been characterized in vitro. At stoichiometric levels rSir3p interacts with intact nucleosomal arrays, mononucleosomes, and naked DNA, as evidenced by formation of supershifted species on native agarose gels. Proteolytic removal of the core histone tail domains inhibits but does not completely abolish rSir3p binding to nucleosomal arrays. The linker DNA in the supershifted complexes remains freely accessible to restriction endonuclease digestion, suggesting that both the tail domains and nucleosomal DNA contribute to rSir3p--chromatin interactions. Together these data indicate that rSir3p cross-links individual nucleosomal arrays into supramolecular assemblies whose physical properties transcend those of typical 10-nm and 30-nm fibers. Based on these data we hypothesize that Sir3p functions, at least in part, by mediating reorganization of the canonical chromatin fiber into functionally specialized higher order chromosomal domains.

A role for a replicator dominance mechanism in silencing.

The role of the natural HMR-E silencer in modulating replication initiation and silencing by the origin recognition complex (ORC) was examined. When natural HMR-E was the only silencer controlling HMR, the silencer's ORC-binding site (ACS) was dispensable for replication initiation but essential for silencing, indicating that a non-silencer chromosomal replicator(s) existed in close proximity to the silencer. Further analysis revealed that regions flanking both sides of HMR-E contained replicators. In contrast to replication initiation by the intact silencer, initiation by the non-silencer replicator(s) was abolished in an orc2-1 mutant, indicating that these replicators were extremely sensitive to defects in ORC. Remarkably, the activity of one of the non-silencer replicators correlated with reduced silencing; inactivation of these replicators caused by either the orc2-1 mutation or the deletion of flanking sequences enhanced silencing. These data were consistent with a role for the ORC bound to the HMR-E silencer ACS in suppressing the function of neighboring ORC molecules capable of inhibiting silencing, and indicated that differences in ORC-binding sites within HMR itself had profound effects on ORC function. Moreover, replication initiation by natural HMR-E was inefficient, suggesting that closely spaced replicators within HMR contributed to an inhibition of replication initiation.

An environmental and respiratory health survey of workers in a grain mill in the Johannesburg area, South Africa.

A respiratory health survey was conducted in a grain mill and the relationship of health indicators to quantitative measures of airborne dust, fungal, and bacterial contamination was examined. Respiratory symptoms were more prevalent in the high dust exposure categories; lung function levels were also higher in the high dust exposure categories, consistent with a "healthy" worker effect. Workers in the three higher dust exposure categories showed either no change or a decrease in lung function over the working week, while workers in the low exposure category demonstrated an improvement in lung function over the working week. Total dust and microbiological (fungal and bacterial) load were found to be significantly related to each other, and the relationship of microbiological load to lung function level and changes over the working week were similar to those found for total dust.

Relationship of respiratory health status to grain dust in a Witwatersrand grain mill: comparison of workers' exposure assessments with industrial hygiene survey findings.

Objective measures of exposure furnished by dust monitoring are both costly and time consuming and require a sufficient level of technology. However, they are important in demonstrating exposure-response relationships, in furnishing information necessary to establish environmental control levels, and to assess if interventions, for instance, improving dust control, have been effective. In this paper respiratory symptoms and cross-shift changes in spirometric lung function were related to dust exposure level in a grain mill assessed in two ways, subjectively (by workers themselves on a four point scale) and objectively (by personal dust monitoring). Health indicators that depend on the individual's perception (e.g., symptoms) correlated more closely with the subjectively assessed dust category, while health indicators that were measured objectively (e.g., cross-week FVC and FEV1 change) correlated more closely with the objectively assessed dust category. However, the patterns of relationship of respiratory health indicators to either dust category were similar, and exposure assessed by one method was, to a large extent, a proxy for the other. The most significant predictor of workers' choice of dust exposure category was the measured dust level. These findings indicate that exposure categories based on workers' assessment of dustiness can be a useful tool in etiologic research, in particular in establishing exposure-response relationships.

Biosynthesis of human acute-phase serum amyloid A protein (A-SAA) in vitro: the roles of mRNA accumulation, poly(A) tail shortening and translational efficiency.

Human 'acute-phase' serum amyloid A protein (A-SAA) is a major acute-phase reactant (APR) and an apolipoprotein of high density lipoprotein 3 (HDL3). We have examined several parameters of A-SAA biosynthesis in PLC/PRF/5 hepatoma cells in response to monocyte conditioned medium (MoCM) and dual treatment with interleukin-1 beta and interleukin-6 (IL-1 beta + IL-6). Treatment of PLC/PRF/5 cells with MoCM or IL-1 beta + IL-6 caused a dramatic and rapid increase in A-SAA mRNA and protein synthesis; A-SAA mRNA was first detectable at 3 h, with peak levels reached by 24 h. A-SAA mRNA accumulation is accompanied by a gradual and homogeneous decrease in the length of the A-SAA poly(A) tail; the poly(A) tail shortening does not apparently affect the intrinsic stability of A-SAA mRNA. Analysis of RNA isolated from the ribonucleoprotein, monosome and polysome fractions of cytokine-treated PLC/PRF/5 cells showed that most A-SAA mRNA was associated with small polyribosomes, regardless of time post-stimulus, suggesting that the translational efficiency of A-SAA mRNA is constant throughout cytokine-driven induction. Moreover, the transit time of A-SAA protein out of the cell is also constant throughout the time course of induction. These data provide evidence of a paradox with regard to the transcriptional upregulation of A-SAA by IL-1 beta + IL-6 and the relative synthesis of A-SAA protein and suggest a role for post-transcriptional control of A-SAA biosynthesis during the acute phase.

Dog serum amyloid A protein. Identification of multiple isoforms defined by cDNA and protein analyses.

Five distinct serum amyloid A (SAA) cDNA clones have been isolated from a library constructed using hepatic mRNA isolated from an individual beagle dog with canine pain syndrome. This implies the existence of at least three SAA genes in the dog genome. One clone predicts a truncated "amyloid A-like" SAA molecule and offers a possible alternative mechanism for the pathogenesis of secondary amyloidosis. Relative to the human and mouse SAA proteins, an additional peptide of eight amino acids is specified by each of the dog cDNA clones. The existence of this peptide in all acute phase dog SAA proteins was confirmed by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate of acute phase high density lipoprotein and provides supporting evidence for gene conversion as a mechanism for maintaining the homogeneity of the SAA gene family within a species. Analysis of hepatic RNA following induction of an acute phase response shows a dramatic increase in SAA mRNA concentration; the SAA transcripts show a transient increase in size early in inflammation due to an increase in polyadenylation.

Major acute-phase reactant synthesis during chronic inflammation in amyloid-susceptible and -resistant mouse strains.

Hepatic levels of mRNA specific for total serum amyloid A (SAA), the SAA1 and SAA2 isotypes, serum amyloid P (SAP), C-reactive protein (CRP), and fibronectin, as well as the plasma concentrations of SAA and SAP were examined in amyloid-resistant (A/J) and amyloid-susceptible (CBA/J) mice during azocasein-induced chronic inflammation. In both strains hepatic SAA and SAP mRNA levels and plasma SAA and SAP protein concentrations increased dramatically during the early stages of inflammation; this was followed by a decrease to concentrations that were maintained at levels considerably higher than background. The ratios of SAA1 and SAA2 mRNA and plasma protein were 1:1 throughout. This indicated that there was no preferential accumulation of mRNA specifying a particular isotype and no preferential synthesis or clearance of a particular isotype during chronic inflammation and the early stages of amyloidogenesis in either strain. Similarly, hepatic SAP mRNA levels in both strains increased dramatically during the early stages of inflammation and were subsequently maintained at elevated levels. Plasma SAP concentrations increased rapidly during the first three days of the study in both A/J and CBA/J mice; however, during the later stages of inflammation, A/J plasma SAP levels decreased to a steady-state concentration that was approximately half that observed in CBA/J mice. Our results identify differences in the hepatic mRNA and plasma protein levels of the major mouse acute-phase reactants (APR) in the amyloid-resistant A/J and amyloid-susceptible CBA/J mouse strains. These findings are consistent with circulating inflammatory APR concentrations contributing, together with other factors, to the onset and pathogenesis of secondary amyloidosis.