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[Determination of aconitine in biological samples].

The determination of aconitine in biological samples by using TLC, HPLC and animal tests is described. TLC is simple, easy to manipulate and highly sensitive. The minimum detection limit was 0.3 microgram, so it can be used as a preliminary screening method. While detecting by HPLC, the characteristic UV-absorption spectrum of aconitine and animal tests were used as the qualitative methods. The characteristic UV absorption wavelengths were (228 +/- 2) nm and (275 +/- 2) nm. The linear range of calibration curve was 2 mg/L-50 mg/L. The coefficient of variation was 0.9996. It is suitable for applications in forensic field.

Aconitine↗

Comparative study of the genomic organization of DNA repeats within the 5'-flanking region of the natural resistance-associated macrophage protein gene (NRAMP1) between humans and great apes.

The human NRAMP1 gene located on Chromosome (Chr) region 2q35 is a candidate gene for increased risk of infection by several intracellular macrophage parasites, including M. tuberculosis and M. leprae. In search for a possible mutational hot spot, we have analyzed a 3.5-kb region 5' to NRAMP1 that is highly enriched for DNA repeat sequences. The repeat sequences could be grouped into one Mer element and six Alu elements, representing five Alu subfamilies, that had integrated in the same DNA region during successive rounds of Alu retropositional activity. Comparative sequence analysis of the Alu cluster region in humans, chimpanzee (Pan paniscus), and gorilla (Gorilla gorilla) revealed only modest sequence variability and failed to detect any evidence for genomic instability of the highly repetitive DNA region. These results show that sequence length variants in the Alu-flanking regions as well as nucleotide substitutions are the most common genomic variations even in a region of extreme Alu-clustering. Moreover, the high degree of sequence conservation among three primate species argues against the Alu cluster being the site of frequent genomic rearrangements or other frequent genetic events that might influence NRAMP1 expression.

Animals↗

Chromosomes and spermatozoa of the African great apes.

We have analysed the chromosome constitution of 8 chimpanzees (Pan troglodytes), 3 pygmy chimpanzees (Pan paniscus), and 16 gorillas (Gorilla gorilla). In the chimpanzees, the frequency of brilliant Q-band polymorphisms accounted for 8.85 regions per individual, and in the gorilla 14.9, whereas in man it was only 2.9-4.6. Variation in the amount of constitutive heterochromatin was also observed, and the gorilla appeared to be the most variable of all. A detailed analysis of the spermatozoa of the African apes showed that the gorilla produced pleiomorphic spermatozoa. As the animals under investigation were phenotypically normal, of proven fertility, and were kept in unheated cages, it is probable that gorillas, like human males, normally exhibit pleiomorphic spermatozoa. This makes it unlikely that clothing-induced hyperthermia could account for pleiomorphic spermatozoa in man. The modal cell types in the ejaculates of men and gorillas were also morphologically identical. The less frequent cell types defined as morphologically abnormal spermatozoa were also very similar and occurred in similar proportions. It is therefore impossible to distinguish between man and the gorilla by a simple examination of the ejaculate, although it is possible to distinguish between man and the chimpanzees or between the gorilla and the chimpanzees. Both species of chimpanzee produced identical spermatozoa. With quinacrine fluorescence, F-bodies were observed in the sperm head of the chimpanzees and the gorilla. In the chimpanzee and the pygmy chimpanzee, F-bodies correspond to brilliant autosomal regions, whereas in the gorilla they probably correspond both the the autosomes and the Y chromosome. There was no consistently visible F-body in the sperm head of the chimpanzee or the gorilla; in the pygmy chimpanzee, one fluorescent region appeared, and it probably corresponded to a brilliant region in autosomal pair No. 23.

Animals↗

The complete nucleotide sequence of a pre-core mutant of hepatitis B virus implicated in fulminant hepatitis and its biological characterization in chimpanzees.

Hepatitis B virus (HBV) with an in-frame stop codon within the pre-core region of the virus genome caused fulminant hepatitis in two individuals. Serum from a chronically infected patient who was the source of the virus was inoculated into three chimpanzees at dilutions of 10(-1), 10(-3), and 10(-7). All three chimpanzees developed acute hepatitis B with relatively high peak values of liver enzymes in their serum. The complete nucleotide sequence of virus DNA recovered from the chimpanzee serum by enzymatic amplification was identical with that from the human serum. By comparing the sequence of this strain (HT) with that of 32 published HBV genomes, changes in nucleotides and predicted amino acids that were rarely or never found in other HBV isolates were identified. Thirteen such nucleotides were found within the cis-acting regulatory elements, of which 6 were within the enhancer II-core promoter region. Twenty-four rare or unique changes in amino acids were found in open reading frames, of which 15 occurred in the region that spanned the 3' half of the X gene, through the pre-core/core gene, to the 5' end of the polymerase gene. Thus, an HBV pre-core stop mutant implicated in fulminant hepatitis is highly infectious, induces severe hepatitis in chimpanzees, and possesses significant genetic variation from reported HBV isolates.

Animals↗

A prevalent POLG CAG microsatellite length allele in humans and African great apes.

The human nuclear gene for the catalytic subunit of mitochondrial DNA polymerase gamma ( POLG) contains within its coding region a CAG microsatellite encoding a polyglutamine repeat. Previous studies demonstrated an association between length variation at this repeat and male infertility, suggesting a mechanism whereby the prevalent (CAG)(10) allele, which occurs at a frequency of >80% in different populations, could be maintained by selection. Sequence analysis of the POLG CAG microsatellite region of more than 1000 human chromosomes reveals that virtually all allelic variation at the locus is accounted for by length variation of the CAG repeat. Analysis of POLG from African great apes shows that a prevalent length allele is present in each species, although its exact length is species-specific. In common chimpanzee ( Pan troglodytes) a number of different sequence variants contribute to the prevalent length allele, strongly supporting the idea that the length of the POLG microsatellite region, rather than its exact nucleotide or amino acid sequence, is what is maintained. Analysis of POLG in other primates indicates that the repeat has expanded from a shorter, glutamine-rich sequence, present in the common ancestor of Old and New World monkeys.

Amino Acid Sequence↗

Construction of fingerprinting and genetic diversity of mulberry cultivars in China by ISSR markers.

The ISSR fingerprintings of 24 mulberry cultivars were constructed. Totally 80 bands were produced using 17 primers selected from 20 primers. Of them, 40 bands showed polymorphism. From the bands amplified, there were three independent ways to identify the mulberry varieties, such as unique ISSR markers, unique band patterns and a combination of the band patterns provided by different primers. ISSRs were very effective in differentiating the mulberry varieties. The mean genetic similarity coefficient, the mean Nei's gene diversity (h), and the mean Shannon's Information index (I) of mulberry cultivars were 0.8731, 0.1210, and 0.1942, respectively. This suggests that the genetic diversity of mulberry cultivars was low and the genetic base was narrow. Both UPGMA cluster and PCA (Principal Coordinates Analysis) analysis showed clear genetic relationships among the 24 mulberry cultivars. The major clusters were related to known pedigree relationships.

China↗

V3 sequence analysis and biological characterization of HIV-1 isolates from asymptomatic and early symptomatic Tanzanian individuals.

The aim of this study was to determine HIV-1 V3 sequences, in vitro biological characteristics and co-receptor usage of virus isolates from Tanzania. Virus was isolated from 14 of 17 samples investigated. Four of the isolates induced syncytia in MT-2 cells and used the CXCR4 co-receptor, while the remaining 10 isolates used the CCR5 co-receptor characteristic of non-MT-2 tropic viruses. One of the four MT-2 tropic isolates also used the CCR5 and CCR3 co-receptors. Proviral DNA was detected in all 14 isolates and PCR products were subjected to DNA sequencing. Unambiguous V3 amino acid sequences were obtained from 11 amplificates. Phylogenetic analysis indicated that these sequences were divergent and clustered in HIV-1 subtypes A, C or D. Sequences from the viruses that induced syncytia in MT-2 cells presented characteristic V3 phenotype-associated amino acids. Results of co-receptor analysis are in concordance with the isolate phenotype as determined by replication and induction of syncytia in MT-2 cells. The considerable diversity illustrated by a limited number of isolates from Tanzania is in accordance with reports from other regions of Africa.

Adolescent↗

HIV infections in northwestern Cameroon: identification of HIV type 1 group O and dual HIV type 1 group M and group O infections.

HIV-1 strain diversity was examined in a study population that consisted of hospital and clinic patients from seven cities and villages located in the northwestern regions of Cameroon. Specimens were screened using a serological algorithm designed to identify HIV-1 group M, N, and O, and SIVcpz-like infections followed by RT-PCR amplification to characterize the infecting virus. The results show that the HIV epidemic in northwest Cameroon is dominated by HIV-1 group M CRF02_AG infections (57%). Additional group M subtypes present include A, D, F2, G, and CRF01_AE. Based on discordant subtype classification between gag and env sequences, a high percentage (23%) of viral strains appear to be unique intersubtype recombinants with the majority (88%) involving recombination with CRF02_AG. Group O prevalence is low accounting for only 0.4% of HIV infections. However, group O strain diversity is high; isolates from clades I, IV, and V, as well as unclassified and recombinant strains, were found. Three dual infections by HIV-1 group M and group O were identified and characterized. In two specimens, both group M and O sequences were amplified in gag, pol, and env suggesting the presence of both viruses. Analysis of the third specimen shows the presence of a group O virus and an intergroup M/O recombinant virus. Finally, no infections due to HIV-1 group N or SIVcpz-like strains were found in the study population.

Amino Acid Sequence↗

Likelihood and Bayes estimation of ancestral population sizes in hominoids using data from multiple loci.

Polymorphisms in an ancestral population can cause conflicts between gene trees and the species tree. Such conflicts can be used to estimate ancestral population sizes when data from multiple loci are available. In this article I extend previous work for estimating ancestral population sizes to analyze sequence data from three species under a finite-site nucleotide substitution model. Both maximum-likelihood (ML) and Bayes methods are implemented for joint estimation of the two speciation dates and the two population size parameters. Both methods account for uncertainties in the gene tree due to few informative sites at each locus and make an efficient use of information in the data. The Bayes algorithm using Markov chain Monte Carlo (MCMC) enjoys a computational advantage over ML and also provides a framework for incorporating prior information about the parameters. The methods are applied to a data set of 53 nuclear noncoding contigs from human, chimpanzee, and gorilla published by Chen and Li. Estimates of the effective population size for the common ancestor of humans and chimpanzees by both ML and Bayes methods are approximately 12,000-21,000, comparable to estimates for modern humans, and do not support the notion of a dramatic size reduction in early human populations. Estimates published previously from the same data are several times larger and appear to be biased due to methodological deficiency. The divergence between humans and chimpanzees is dated at approximately 5.2 million years ago and the gorilla divergence 1.1-1.7 million years earlier. The analysis suggests that typical data sets contain useful information about the ancestral population sizes and that it is advantageous to analyze data of several species simultaneously.

Algorithms↗

Antibodies to V3 loop peptides derived from chimpanzee lentiviruses and the divergent HIV-1ANT-70 isolate in human sera from different geographic regions.

OBJECTIVE: To study the spread of antibodies to V3 loop peptides of two chimpanzee lentiviruses and the divergent HIV-1ANT-70 isolate (group O) in human sera from different geographic regions, and to compare this with reactions to peptides from known North American (subtype B) and Zaïrean (subtype D) strains. METHODS: A total 2495 HIV-1-antibody-positive sera from nine countries were tested by enzyme-linked immunosorbent assay for antibodies to the V3 loop of 10 HIV/SIV isolates (including MN, SF2, HXB2, RF, MAL, ELI, Z6 and ANT-70 for HIV-1, and cpz-gab and cpz-ant for SIV). RESULTS: In each country, the highest prevalences were observed against the MN peptide (58.8-91.7%). Seroreactivity to other peptides from subtype B were generally lower. Prevalences of antibodies to V3 peptides derived from Zaïrean strains belonging to subtype D were generally lower than to subtype B. Relative high prevalences of sera reactive with the SIVcpz-gab V3 peptide were observed. The lowest rates were seen in Brazil (4.2%) and Belgium (25.7%). Among the African countries, the prevalence rates varied between 30.1 and 67.6%. Prevalence to the V3 loop derived from the SIVcpz-ant strains was much lower. Prevalence of sera reactive to the ANT-70 V3 loop peptide was very low, and the highest rates were observed in Cameroon (10.2%), Niger (6%) and Gabon (4.6%). Only the sera reactive to the ANT-70 V3 loop peptide from Cameroon and Gabon were confirmed on a specific HIVANT-70 Western blot (i.e., presence of antibodies to the envelope protein gp 120). CONCLUSIONS: The extent to which different V3 peptide reactivity patterns reflect the circulation of different HIV-1 strains in a particular population is not yet clear. However, V3 peptide serology using the very specific V3 peptide of the HIVANT-70 is a good indicator of the very aberrant group O in a particular population.

Amino Acid Sequence↗

Evidence for positive selection at the pantophysin (Pan I) locus in walleye pollock, Theragra chalcogramma.

Nucleotide polymorphism at the pantophysin (Pan I) locus in walleye pollock, Theragra chalcogramma, was examined using DNA sequence data. Two distinct allelic lineages were detected in pollock, resulting from three amino acid replacement mutations in the first intravesicular domain of the protein. The common Pan I allelic group, comprising 94% of the samples, was less polymorphic (pi = 0.005) than the uncommon group (pi = 0.008), and nucleotide diversity in both was higher than for two allelic lineages in the related Atlantic cod, Gadus morhua. Phylogenetic analyses of Pan I sequences from these two species did not clearly resolve orthology among allelic groups, in part because of recombination that has occurred between the two pollock lineages. Conventional tests of neutrality comparing polymorphisms within and between homologous regions of the Pan I locus in walleye pollock and Atlantic cod did not detect the effects of selection. This result is likely attributed to low levels of synonymous divergence among allelic lineages and a lack of mutation-drift equilibrium inferred from nucleotide mismatch frequency distributions. However, the ratio of nonsynonymous to synonymous substitutions per site (dN/dS) exceeded unity in two intravesicular domains of the protein and the influence of positive selection at multiple codon sites was strongly inferred through the use of maximum-likelihood analyses. In addition, the frequency spectrum of linked neutral variation showed indirect effects of adaptive hitchhiking in pollock resulting from a selective sweep of the common allelic lineage. Recombination between the two allelic classes may have prevented complete loss of the older, more polymorphic lineage. The results suggest that recurrent sweeps driven by positive selection is the principle mode of evolution at the Pan I locus in gadid fishes.

Adaptation, Biological↗

Prevalence of dyslipidemia and mean blood lipid values in Taiwan: results from the Nutrition and Health Survey in Taiwan (NAHSIT, 1993-1996).

This paper reports the blood lipid status of people aged 4 years and older in Taiwan. The data is based on the Nutrition and Health Survey in Taiwan (NAHSIT: 1993-1996), which adopted a multi-stage, stratified clustering sampling scheme. Altogether, 5097 subjects (2451 males and 2646 females) had data on triglyceride and 5643 subjects (2736 males and 2907 females) had data on cholesterol. We found that (a) cholesterol levels of males were lower than females in mid-to old age group (> or = 45 years old); (b) triglyceride values of females were lower than males in young adulthood (19-44 years), but higher than males after the age of 45 years, and (c) adult females had higher HDL-C value and lower ratio of total cholesterol to HDL-C than males. The prevalence of hypercholesterolemia was 10.2% in adult males and 12.6% in mid-to-old aged men, and that in females was 11.2% and 24.4%, respectively. The prevalence of hypertriglyceridemia was 13.4% and 6.1% in adult males and females (> or = 19 years as a whole), respectively. It was 12.3% in mid-to-old aged men (> or = 45 years), and 11.9% in women. The mean cholesterol values were similar to values of several previous surveys in different areas of Taiwan. But it was higher than those in some areas of Mainland China, and lower than those of western countries. People in metropolitan cities had a higher level of blood cholesterol than other areas. The average triglyceride values of males and females were higher than those of previous studies in Taiwan and of people in Mainland China. Mountainous stratum with predominantly aboriginal residents had higher level of triglycerides and body mass index (BMI) than other strata. The associations between dietary intakes of men and women and blood lipids were examined controlling for age and BMI. Result showed that Keys score, which was derived from saturated fat, polyunsaturated fat and dietary cholesterol of a 24-hour recall, was positively related to blood cholesterol and LDL-C in men, but not in women. Average alcohol intakes per day were related to HDL-C positively, but LDL-C negatively in men and women. The regional differences in blood lipid profiles in Taiwan are consistent with the dietary and life-style variations island-wide.

Adolescent↗

Histologic examination of bone development in juvenile chimpanzees.

The purpose of this study is to determine whether histologic skeletal development in chimpanzees (Pan troglodytes) differs from that in humans. Currently, minimal quantitative data are available on the bone histology of great apes. In addition to providing baseline data on juvenile chimpanzee bone histology, the data generated by this study have potential applications for studying the comparative development between chimpanzees and humans and other primates, as well as investigating the evolution of human bone development, differences in development among limb elements, and differences in histology related to locomotor function. The study sample includes thin sections from the femoral, tibial, and fibular midshafts of 13 chimpanzees originally prepared by Kerley ([1966] Tulane Stud. Zool. 13:71-82) as part of a study on skeletal age changes in the chimpanzee. Twelve juveniles, ranging in known age from 2-15.3 years, and one adult, with a known age of 35 years, are represented. For each specimen, numbers of osteons, osteon fragments, and non-Haversian canals were counted, and percent lamellar bone was estimated. Results were compared with data extracted from Kerley ([1965] Am. J. Phys. Anthropol. 23:149-164) on a juvenile human sample. Results indicate that juvenile chimpanzees and humans exhibit similar age-related changes in histologic variables. However, age is not as strong a predictor of variation in microstructural variables in chimpanzees as it is in humans.

Animals↗

The opioid receptor-ligand network in human cancers: pan-cancer multi-omics profiling and translational implications.

BACKGROUND: Opioid receptor-ligand signalling has been implicated in tumour biology and perioperative outcomes; however, its pan-cancer molecular landscape and clinical relevance remain incompletely defined. METHODS: We performed a pan-cancer multi-omics analysis of eight predefined opioid receptor-ligand genes across 33 tumour types from The Cancer Genome Atlas. Analyses included gene expression analysis using the linear models for microarray data (limma) package, genomic alterations, DNA methylation, regulatory network inference, pathway activity estimation using gene set variation analysis, and survival modelling. Multivariable Cox regression models were adjusted for age, sex, and tumour stage. RESULTS: Opioid receptor-ligand genes exhibited heterogeneous and generally low-to-moderate expression across tumour types. Genomic and epigenetic alterations were tumour-specific and variably associated with gene expression. Selected genes showed associations with overall survival in a tumour-dependent manner; however, these associations were attenuated after adjustment for clinical covariates and were accompanied by wide confidence intervals in some cohorts. Pathway analyses suggested associations with broader biological programmes, including epithelial-mesenchymal transition and immune-related pathways. Regulatory analyses identified candidate transcription factors and miRNAs, although these findings are exploratory. CONCLUSIONS: This pan-cancer analysis provides a systematic overview of opioid receptor-ligand gene features across human cancers. The observed associations are context-dependent and should be interpreted as hypothesis-generating. Further mechanistic and prospective studies are required to determine the clinical relevance of opioid signalling in cancer and perioperative settings.

Humans↗

Predictors of dietary heterocyclic amine intake in three prospective cohorts.

Cooking meat creates heterocyclic amines (HCAs) through pyrolysis of amino acids and creatinine. Although recognized as mutagenic, the etiological role of HCA in human cancer is unclear, due to the lack of information on the effect of typical food cooking methods on HCA concentrations and on variation in HCA exposure in populations. We estimated overall daily dietary HCA intake and variation in intake between individuals, using recent data on HCA concentrations in various meats prepared by cooking methods, temperatures, and times common in United States in the 1990s. Random samples of 250 participants from each of three large prospective cohorts were mailed a questionnaire to assess frequency of consumption, cooking method, and typical outside appearance of pan-fried, broiled, and grilled or barbecued chicken, fish, hamburger, and steak; fried, microwaved, and broiled bacon; fried sausage; roast beef; and homemade gravy. The 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), and 2-amino-3,4,8-trimethylimidazo[4,5,f]quinoxaline (DiMeIQx) concentrations, measured in composite samples by solid-phase extraction and high-performance liquid chromatography, were assigned to each food, cooking method, and doneness level. The dietary reports showed approximately 30-fold relative variation in 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline intake, 20-fold for 2-amino- -methyl-6-phenylimidazo[4,5-b]pyridine, and over 110-fold for 2-amino-3,4,8-trimethylimidazo[4,5,f]quinoxaline, when the 10th and 90th percentiles of HCA intake were compared (90th/10th percentile value). These reported variations in HCA exposure among participants in these three large cohorts indicates that estimation of HCA intake and determination of association with disease risk are feasible, if additional information on meat cooking methods is obtained.

Adult↗

Preliminary vocal repertoire and vocal communication of wild bonobos (Pan paniscus) at Lilungu (Democratic Republic of Congo).

Vocal communication in bonobos, Pan paniscus, has been studied mainly in captive groups. The few studies carried out in the wild are on only specific aspects of their behaviour. The aim of this study is to give a preliminary description of the vocal repertoire and a qualitative account of the contextual use of the vocalizations described. Observations were carried out on three wild communities, with a total of 68 individuals, for 685 h. No artificial feeding was used at any stage of the study. The vocal repertoire is composed of 15 vocal units and 19 sequences. There is great variability both in the sequences and in the vocal units, and the categories are not always completely discrete. The repertoire is compared to that of captive bonobos described by De Waal. Most sequences are used in various contexts, and for each behaviour there is a complete array of sequences and their variation. The behaviour patterns observed and the sequences uttered by the animals in each context are discussed.

Animals↗

A framework for food-based dietary guidelines in the European Union.

UNLABELLED: The nutrient targets derived from analysis of the relation between nutrient intake and disease prevalence or other scientific evidence, have to be translated into food-based guidelines in order to be understood by the general population. Furthermore, Food-Based Dietary Guidelines (FBGD) have to be realistic, attainable and culturally acceptable and should also give consideration to relevant social, economic, agricultural and environmental factors affecting food availability and eating patterns. This requires a thorough understanding of the relation between foods, food patterns and nutrient intakes in the population. The aims of Working Party 2 were to propose a framework for strategies in the development of FBDG and to examine existing data for nutrient and food intakes in the EU. METHODS: The over-all strategy given by the joint FAO/WHO consultation 1995 was used as the starting point, i.e. target foods or food patterns for public health nutrition programmes should be identified from an analysis of prevailing food and nutrient intakes. Prevailing data for food and nutrient intakes in 14 EU countries were examined and different principles and options for the derivation of FBDG were explored. Methodological issues and their influence on the interpretation of data for the development of FBDG were also examined. RESULTS: The process from nutrients to foods can be briefly: 1) identification of major food sources of the nutrient of interest, 2) identification of foods contributing substantially to population intakes, 3) identification of foods or food patterns compatible with desirable nutrient intakes or explaining variations in nutrient intakes, 4) formulation of FBDG into foods, portion sizes, frequency of intake, meal composition taking attainability and acceptability as well as compatibility of co-existing guidelines into account. The level of complexity that can be applied in the analytical approaches depends on the characteristics of available intake data. A detailed analysis requires data on an individual level for nutrients, foods, food patterns, eating and meal habits etc. When individual data are available different analytical approaches (examination of distribution of intakes, correlation analysis between foods and nutrients, examination of food intakes in compliers/non-compliers to nutrient goals, discriminant analysis, cluster/factor analysis) can be used to identify key foods or food patterns fulfilling nutrient goals. The examination of prevailing food and nutrient intake data in the EU revealed: a number of methodological differences in approaches to dietary surveys exist in the EU countries e.g., regarding methods used, selection of population, classification of foods, which have to be kept in mind in pan-EU comparisons at present there is a substantial gap between actual intakes and present nutrient goals suggesting that major changes of dietary habits are needed while some food patterns were consistently related to intake of specific nutrients in most EU countries, other patterns showed large variations between countries methodological issues, such as survey duration, survey techniques, under- or over-reporting, could have substantial influence on the identification of target foods or food patterns. CONCLUSIONS: A science-based analysis of nutrient and food intakes allows development of FBDG, which, if implemented, are likely to result in mean population intakes closer to nutrient goals. Acknowledging the social and cultural differences within the EU as well as the need to focus on the most relevant public health problem in the population, FBDG should first be developed within member states. Harmonisation of survey methods within the EU would facilitate development of regional and EU FBDG.

Cluster Analysis↗