Apolipoprotein C Polymorphism in Sheep: Allele Frequencies and Association with Plasma Lipid and Lipoprotein Levels

The genetic polymorphism of apolipoprotein C in normal plasma of four European sheep breeds (Suffolk, Corriedale, Cheviot, and Finn) was first detected using one-dimensional polyacrylamide gel isoelectric focusing (pH 2.5–5.0) followed by immunoblotting with antihuman apolipoprotein CII antibody. Six phenotypes (1-1, 2-1, 2-2, 3-1, 3-2, and 3-3) were identified in the 4.3–4.8 pH range, consisting of the combination of three isoform groups. On the basis of family and population data, these phenotypes were controlled autosomally by three codominant alleles, designated APOC*1, APOC*2, and APOC*3, the first being the most common allele. The frequency distributions of these alleles were similar between the Suffolk and Corriedale sheep, and between the Cheviot and Finn sheep. The former breeds had a significantly lower APOC*2 frequency than the latter breeds (P< 0.001). The mean plasma total-, HDL- and LDL-cholesterol levels of type 3-1 animals were significantly higher compared to type 1-1 animals in the Suffolk sheep (P 0.04). However, these differences were not seen in the Corriedale sheep     

A method for detecting rash and fever illness‐associated viruses using multiplex reverse transcription polymerase chain reaction

In this study, a new multiplex RT‐PCR method for detecting various viral genes in patients with rash and fever illnesses (RFIs ) was constructed. New primer sets were designed for detection of herpes simplex viruses 1 and 2 (HSV1 and 2), and Epstein–Barr virus (EBV). The newly designed and previously reported primer sets were used to detect 13 types of RFI‐associated viruses by multiplex RT‐PCR assay systems. Moreover, to eliminate non‐specific PCR products, a double‐stranded specific DNase was used to digest double‐stranded DNA derived from the templates in clinical specimens. RFI‐associated viruses were detected in 77.0% of the patients (97/126 cases) by the presented method, multiple viruses being identified in 27.8% of the described cases (35/126 cases). Detected viruses and clinical diagnoses were compatible in 32.5% of the patients (41/126 cases). Sensitivity limits for these viruses were estimated to be 10¹–10³ copies/assay. Furthermore, non‐specific PCR products were eliminated by a double‐stranded specific DNase with no influence on sensitivity. These results suggest that this method can detect various RFI‐associated viruses in clinical specimens with high sensitivity and specificity.

     

Patterns of plant diversity at high altitudes on the Qinghai-Tibetan Plateau

Aims To describe the biodiversity patterns of plants along an altitudinal gradient on the Qinghai-Tibetan Plateau and to clarify the bias in plant specimen records at high altitude.Methods We conducted a large-scale investigation of vegetation at a wide range of altitudes, focusing on a high-altitudinal range (3?200–5?200 m) at different locations on the Qinghai-Tibetan Plateau. We then compared the altitudinal distribution of plant species obtained from our field investigation with that in plant specimen records from published sources and an online database.Important findings Our data provide evidence that altitude plays a large role in regulating species composition on the Qinghai-Tibetan Plateau. We could not, however, detect a clear relationship between altitude and species richness, although a weak monotonically increasing trend of richness was detected with increasing altitude. According to specimen records, most species have been sampled at a wide range of altitudes, and the average range of 145 species is>2?000 m. Despite this wide range, more than half of the species we observed were at higher altitudes than the specimen records indicate. High-altitude areas have probably been so poorly sampled that only a small fraction of the resident species has been recorded. This study clearly shows the regional bias of specimen records in the Qinghai-Tibetan Plateau.     

Genetic analyses of the functions of [NiFe]-hydrogenase maturation endopeptidases in the hyperthermophilic archaeon Thermococcus kodakarensis

The maturation of [NiFe]-hydrogenases requires a number of accessory proteins, which include hydrogenase-specific endopeptidases. The endopeptidases carry out the final cleavage reaction of the C-terminal regions of [NiFe]-hydrogenase large subunit precursors. The hyperthermophilic archaeon Thermococcus kodakarensis harbors two [NiFe]-hydrogenases, a cytoplasmic Hyh and a membrane-bound Mbh, along with two putative hydrogenase-specific endopeptidase genes. In this study, we carried out a genetic examination on the two endopeptidase genes, TK2004 and TK2066. Disruption of TK2004 resulted in a strain that could not grow under conditions requiring hydrogen evolution. The Mbh large subunit precursor (pre-MbhL) in this strain was not processed at all whereas Hyh cleavage was not affected. On the other hand, disruption of TK2066 did not affect the growth of T. kodakarensis under the conditions examined. Cleavage of the Hyh large subunit precursor (pre-HyhL) was impaired, but could be observed to some extent. In a strain lacking both TK2004 and TK2066, cleavage of pre-HyhL could not be observed. Our results indicate that pre-MbhL cleavage is carried out solely by the endopeptidase encoded by TK2004. Pre-HyhL cleavage is mainly carried out by TK2066, but TK2004 can also play a minor role in this cleavage.

     

Characterization of newly developed SSLP markers for the rat

We have isolated more than 12,000 clones containing microsatellite sequences, mainly consisting of (CA)n dinucleotide repeats, using genomic DNA from the BN strain of laboratory rat. Data trimming yielded 9636 non-redundant microsatellite sequences, and we designed oligonucleotide primer pairs to amplify 8189 of these. PCR amplification of genomic DNA from five different rat strains yielded clean amplification products for 7040 of these simple-sequence-length-polymorphism (SSLP) markers; 3019 markers had been mapped previously by radiation hybrid (RH) mapping methods (Nat Genet 22, 27–36, 1998). Here we report the characterization of these newly developed microsatellite markers as well as the release of previously unpublished microsatellite marker information. In addition, we have constructed a genome-wide linkage map of 515 markers, 204 of which are derived from our new collection, by genotyping 48 F₂ progeny of (OLETFxBN)F₂ crosses. This map spans 1830.9 cM, with an average spacing of 3.56 cM. Together with our ongoing project of preparing a whole-genome radiation hybrid map for the rat, this dense linkage map should provide a valuable resource for genetic studies in this model species. Received: 8 July 1999 / Accepted: 3 December 1999     

Characterization of the Gut Microbiota of Papua New Guineans Using Reverse Transcription Quantitative PCR

There has been considerable interest in composition of gut microbiota in recent years, leading to a better understanding of the role the gut microbiota plays in health and disease. Most studies have been limited in their geographical and socioeconomic diversity to high-income settings, and have been conducted using small sample sizes. To date, few analyses have been conducted in low-income settings, where a better understanding of the gut microbiome could lead to the greatest return in terms of health benefits. Here, we have used quantitative real-time polymerase chain reaction targeting dominant and sub-dominant groups of microorganisms associated with human gut microbiome in 115 people living a subsistence lifestyle in rural areas of Papua New Guinea. Quantification of Clostridium coccoides group, C. leptum subgroup, C. perfringens, Bacteroides fragilis group, Bifidobacterium, Atopobium cluster, Prevotella, Enterobacteriaceae, Enterococcus, Staphylococcus, and Lactobacillus spp. was conducted. Principle coordinates analysis (PCoA) revealed two dimensions with Prevotella, clostridia, Atopobium, Enterobacteriaceae, Enterococcus and Staphylococcus grouping in one dimension, while B. fragilis, Bifidobacterium and Lactobacillus grouping in the second dimension. Highland people had higher numbers of most groups of bacteria detected, and this is likely a key factor for the differences revealed by PCoA between highland and lowland study participants. Age and sex were not major determinants in microbial population composition. The study demonstrates a gut microbial composition with some similarities to those observed in other low-income settings where traditional diets are consumed, which have previously been suggested to favor energy extraction from a carbohydrate rich diet.