HDL subspecies : association with low HDL-C, obesity, and metabolic syndrome

AIMS An independent, powerful coronary heart disease (CHD) predictor is a low level of high-density lipoprotein cholesterol (HDL-C). Discoidal preβ-HDL particles and large HDL2 particles are the primary cholesterol acceptors in reverse cholesterol transport, a key anti-atherogenic HDL mechanism. The quality of HDL subspecies may provide better markers of HDL functionality than does HDL-C alone. We aimed I) to study whether alterations in the HDL subspecies profile exist in low-HDL-C subjects II) to explore the relationship of any changes in HDL subspecies profile in relation to atherosclerosis and metabolic syndrome; III) to elucidate the impact of genetics and acquired obesity on HDL subspecies distribution. SUBJECTS The study consisted of 3 cohorts: A) Finnish families with low HDL-C and premature CHD (Study I: 67 subjects with familial low HDL-C and 64 controls; Study II: 83 subjects with familial low HDL-C, 65 family members with normal HDL-C, and 133 controls); B) a cohort of 113 low- and 133 high-HDL-C subjects from the Health 2000 Health Examination Survey carried out in Finland (Study III); and C) a Finnish cohort of healthy young adult twins (52 monozygotic and 89 dizygotic pairs) (Study IV). RESULTS AND CONCLUSIONS The subjects with familial low HDL-C had a lower preβ-HDL concentration than did controls, and the low-HDL-C subjects displayed a dramatic reduction (50-70%) in the proportion of large HDL2b particles. The subjects with familial low HDL-C had increased carotid atherosclerosis measured as intima-media-thickness (IMT), and HDL2b particles correlated negatively with IMT. The reduction in both key cholesterol acceptors, preβ-HDL and HDL2 particles, supports the concept of impaired reverse cholesterol transport contributing to the higher CHD risk in low-HDL-C subjects. The family members with normal HDL-C and the young adult twins with acquired obesity showed a reduction in large HDL2 particles and an increase in small HDL3 particles, which may be the first changes leading to the lowering of HDL-C. The low-HDL-C subjects had a higher serum apolipoprotein E (apoE) concentration, which correlated positively with the metabolic syndrome components (waist circumference, TG, and glucose), highlighting the need for a better understanding of apoE metabolism in human atherosclerosis. In the twin study, the increase in small HDL3b particles was associated with obesity independent of genetic effects. The heritability estimate, of 73% for HDL-C and 46 to 63% for HDL subspecies, however, demonstrated a strong genetic influence. These results suggest that the relationship between obesity and lipoproteins depends on different elements in each subject. Finally, instead of merely elevating HDL-C, large HDL2 particles and discoidal preβ-HDL particles may provide beneficial targets for HDL-targeted therapy.

Foodborne Yersinia : Identification and Molecular Epidemiology of Isolates from Human Infections

Yersinia enterocolitica and Yersinia pseudotuberculosis are among the major enteropathogenic bacteria causing infections in humans in many industrialized countries. In Finland, Y. pseudotuberculosis has caused 10 outbreaks among humans during 1997-2008. Some of these outbreaks have been very extensive involving over 400 cases; mainly children attending schools and day-care. Y. enterocolitica, on the contrary, has caused mainly a large number of sporadic human infections in Finland. Y. pseudotuberculosis is widespread in nature, causing infections in a variety of domestic and wild animals. Foodborne transmission of human infections has long been suspected, however, attempts to trace the pathogen have been unsuccessful before this study that epidemiologically linked Y. pseudotuberculosis to a specific food item. Furthermore, due to modern food distribution systems, foodborne outbreaks usually involve many geographically separate infection clusters difficult to identify as part of the same outbreak. Among pathogenic Y. enterocolitica, the global predominance of one genetically homogeneous type (bioserotype 4/O:3) is a challenge to the development of genetic typing methods discriminatory enough for epidemiological purposes, for example, for tracing back to the sources of infections. Furthermore, the diagnostics of Y. enterocolitica infections is hampered because clinical laboratories easily misidentify some other members of the Yersinia species (Y. enterocolitica–like species) as Y. enterocolitica. This results in misleading information on the prevalence and clinical significance of various Yersinia isolates. The aim of this study was to develop and optimize molecular typing methods to be used in epidemiological investigations of Y. enterocolitica and Y. pseudotuberculosis, particularly in active surveillance and outbreak investigations of Y. pseudotuberculosis isolates. The aim was also to develop a simplified set of phenotypic tests that could be used in routine diagnostic laboratories for the correct identification of Y. enterocolitica and Y. enterocolitica –like species. A PFGE method designed here for typing of Y. pseudotuberculosis was efficient in linking the geographically dispersed and apparently unrelated Y. pseudotuberculosis infections as parts of the same outbreak. It proved to be useful in active laboratory-based surveillance of Y. pseudotuberculosis outbreaks. Throughout the study period, information about the diversity of genotypes among outbreak and non-outbreak related strains of human origin was obtained. Also, to our knowledge, this was the first study to epidemiologically link a Y. pseudotuberculosis outbreak of human illnesses to a specific food item, iceberg lettuce. A novel epidemiological typing method based on the use of a repeated genomic region (YeO:3RS) as a probe was developed for the detection and differentiation between strains of Y. enterocolitica subspecies palearctica. This method was able to increase the discrimination in a set of 106 previously PFGE typed Finnish Y. enterocolitica bioserotype 4/O:3 strains among which two main PFGE genotypes had prevailed. The developed simplified method was a more reliable tool than the commercially available biochemical test kits for differentiation between Y. enterocolitica and Y. enterocolitica –like species. In Finland, the methods developed for Y. enterocolitica and Y. pseudotuberculosis have been used to improve the identification protocols and in subsequent outbreak investigations.

A systematic-ecological approach to Baltic Sea ice studies of algae and protists

The seasonal occurrence of sea ice that annually covers almost half the Baltic Sea area provides a unique habitat for halo- and cold temperature-tolerant extremophiles. Baltic Sea ice biology has more than 100 years of tradition that began with the floristic observation of species by the early pioneers using light microscopic techniques that were the only thing available at the time. Since the discovery of life within sea ice, more technologies have become available for taxonomy. Electron microscopy and genetic evidence have been used to identify sea ice biota revealing increased numbers of taxa. Meanwhile ecologists have used light microscopic cell enumeration in addition to the chemical and physical properties of sea ice in attempts to explain the food web structure of sea ice and its functions. Thus, during the Baltic winter, the sea ice hosts more abundant and diverse microbial communities than the water column beneath it. These communities are typically dominated by autotrophic diatoms together with a diverse assortment of dinoflagellates, auto- and heterotrophic flagellates, ciliates, metazoan rotifers and bacteria, which are mostly responsible for the recycling of nutrients. This thesis comprises ecological and systematic studies. In addition to the results of the previous studies carried out on landfast ice, the data presented here provide new insight into the spatial distribution of pelagial sea ice, which has remained largely unexplored. The studies reveal spatial heterogeneity in the pelagial sea ice of the Gulf of Bothnia. There were mismatches in chlorophyll-a concentrations and in photosynthetic efficiencies of the communities studied. The temporal succession was followed and experimental studies performed investigating the community responses towards increased or decreased light in landfast ice in the Gulf of Finland. The systematic studies carried out with established dinoflagellate cultures revealed a new resting cyst belonging to common sea ice dinoflagellate, Scrippsiella hangoei (Schiller) Larsen 1995. The cyst can be used to explain the overwintering of this species during prolonged periods of darkness. The dissimilarities and similarities in the material isolated from the sea ice called for description of a new subspecies Heterocapsa arctica ssp. frigida. The cells obtained in the cultured material were unlike those of the previously described species, necessitating description of ssp. frigida. As a result of its own unique habitus, the subspecies had been noted by Finnish taxonomists during the past three decades and thus its annual occurrence and geographical distribution in the Baltic Sea. This illustrates how combining ecology and systematics increases our understanding of organisms.

Phylogeography and hybrid swarms: history of brackish water bivalve diversity in North European marginal seas

This study addressed the large-scale molecular zoogeography in two brackish water bivalve molluscs, Macoma balthica and Cerastoderma glaucum, and genetic signatures of the postglacial colonization of Northern Europe by them. The traditional view poses that M. balthica in the Baltic, White and Barents seas (i.e. marginal seas) represent direct postglacial descendants of the adjacent Northeast Atlantic populations, but this has recently been challenged by observations of close genetic affinities between these marginal populations and those of the Northeast Pacific. The primary aim of the thesis was to verify, quantify and characterize the Pacific genetic contribution across North European populations of M. balthica and to resolve the phylogeographic histories of the two bivalve taxa in range-wide studies using information from mitochondrial DNA (mtDNA) and nuclear allozyme polymorphisms. The presence of recent Pacific genetic influence in M. balthica of the Baltic, White and Barents seas, along with an Atlantic element, was confirmed by mtDNA sequence data. On a broader temporal and geographical scale, altogether four independent trans-Arctic invasions of Macoma from the Pacific since the Miocene seem to have been involved in generating the current North Atlantic lineage diversity. The latest trans-Arctic invasion that affected the current Baltic, White and Barents Sea populations probably took place in the early post-glacial. The nuclear genetic compositions of these marginal sea populations are intermediate between those of pure Pacific and Atlantic subspecies. In the marginal sea populations of mixed ancestry (Barents, White and Northern Baltic seas), the Pacific and Atlantic components are now randomly associated in the genomes of individual clams, which indicates both pervasive historical interbreeding between the previously long-isolated lineages (subspecies), and current isolation of these populations from the adjacent pure Atlantic populations. These mixed populations can be characterized as self-supporting hybrid swarms, and they arguably represent the most extensive marine animal hybrid swarms so far documented. Each of the three swarms still has a distinct genetic composition, and the relative Pacific contributions vary from 30 to 90 % in local populations. This diversity highlights the potential of introgressive hybridization to rapidly give rise to new evolutionarily and ecologically significant units in the marine realm. In the south of the Danish straits and in the Southern Baltic Sea, a broad genetic transition zone links the pure North Sea subspecies M. balthica rubra to the inner Baltic hybrid swarm, which has about 60 % of Pacific contribution in its genome. This transition zone has no regular smooth clinal structure, but its populations show strong genotypic disequilibria typical of a hybrid zone maintained by the interplay of selection and gene flow by dispersing pelagic larvae. The structure of the genetic transition is partly in line with features of Baltic water circulation and salinity stratification, with greater penetration of Atlantic genes on the Baltic south coast and in deeper water populations. In all, the scenarios of historical isolation and secondary contact that arise from the phylogeographic studies of both Macoma and Cerastoderma shed light to the more general but enigmatic patterns seen in marine phylogeography, where deep genetic breaks are often seen in species with high dispersal potential.

Molecular Tuning of Rhodopsins for High Visual Sensitivity in Different Light Environments : Variation in Absorbance Spectrum and Opsin Sequence within and between Species

Visual pigments of different animal species must have evolved at some stage to match the prevailing light environments, since all visual functions depend on their ability to absorb available photons and transduce the event into a reliable neural signal. There is a large literature on correlation between the light environment and spectral sensitivity between different fish species. However, little work has been done on evolutionary adaptation between separated populations within species. More generally, little is known about the rate of evolutionary adaptation to changing spectral environments. The objective of this thesis is to illuminate the constraints under which the evolutionary tuning of visual pigments works as evident in: scope, tempo, available molecular routes, and signal/noise trade-offs. Aquatic environments offer Nature s own laboratories for research on visual pigment properties, as naturally occurring light environments offer an enormous range of variation in both spectral composition and intensity. The present thesis focuses on the visual pigments that serve dim-light vision in two groups of model species, teleost fishes and mysid crustaceans. The geographical emphasis is in the brackish Baltic Sea area with its well-known postglacial isolation history and its aquatic fauna of both marine and fresh-water origin. The absorbance spectrum of the (single) dim-light visual pigment were recorded by microspectrophotometry (MSP) in single rods of 26 fish species and single rhabdoms of 8 opossum shrimp populations of the genus Mysis inhabiting marine, brackish or freshwater environments. Additionally, spectral sensitivity was determined from six Mysis populations by electroretinogram (ERG) recording. The rod opsin gene was sequenced in individuals of four allopatric populations of the sand goby (Pomatoschistus minutus). Rod opsins of two other goby species were investigated as outgroups for comparison. Rod absorbance spectra of the Baltic subspecies or populations of the primarily marine species herring (Clupea harengus membras), sand goby (P. minutus), and flounder (Platichthys flesus) were long-wavelength-shifted compared to their marine populations. The spectral shifts are consistent with adaptation for improved quantum catch (QC) as well as improved signal-to-noise ratio (SNR) of vision in the Baltic light environment. Since the chromophore of the pigment was pure A1 in all cases, this has apparently been achieved by evolutionary tuning of the opsin visual pigment. By contrast, no opsin-based differences were evident between lake and sea populations of species of fresh-water origin, which can tune their pigment by varying chromophore ratios. A more detailed analysis of differences in absorbance spectra and opsin sequence between and within populations was conducted using the sand goby as model species. Four allopatric populations from the Baltic Sea (B), Swedish west coast (S), English Channel (E), and Adriatic Sea (A) were examined. Rod absorbance spectra, characterized by the wavelength of maximum absorbance (λmax), differed between populations and correlated with differences in the spectral light transmission of the respective water bodies. The greatest λmax shift as well as the greatest opsin sequence difference was between the Baltic and the Adriatic populations. The significant within-population variation of the Baltic λmax values (506-511 nm) was analyzed on the level of individuals and was shown to correlate well with opsin sequence substitutions. The sequences of individuals with λmax at shorter wavelengths were identical to that of the Swedish population, whereas those with λmax at longer wavelengths additionally had substitution F261F/Y in the sixth transmembrane helix of the protein. This substitution (Y261) was also present in the Baltic common gobies and is known to redshift spectra. The tuning mechanism of the long-wavelength type Baltic sand gobies is assumed to be the co-expression of F261 and Y261 in all rods to produce ≈ 5 nm redshift. The polymorphism of the Baltic sand goby population possibly indicates ambiguous selection pressures in the Baltic Sea. The visual pigments of all lake populations of the opossum shrimp (Mysis relicta) were red-shifted by 25 nm compared with all Baltic Sea populations. This is calculated to confer a significant advantage in both QC and SNR in many humus-rich lakes with reddish water. Since only A2 chromophore was present, the differences obviously reflect evolutionary tuning of the visual protein, the opsin. The changes have occurred within the ca. 9000 years that the lakes have been isolated from the Sea after the most recent glaciation. At present, it seems that the mechanism explaining the spectral differences between lake and sea populations is not an amino acid substitution at any other conventional tuning site, but the mechanism is yet to be found.