From the Alps to the Mediterranean and beyond: genetics, environment, culture and the "impossible beauty" of Italy.

Since prehistoric times, Italy has represented a bridge between peoples, genes and cultures. Its peculiar geographical position explains why: it is located in the center of the Mediterranean Sea, flanked by the Balkans and the Hellenic Peninsula to the east, Iberia to the west and surrounded by North Africa to the south and central Europe to the north. This makes Italy of extraordinary interest for the study of some different aspects of human diversity. Here we overview current knowledge regarding the relationships between the structure of the genetic variation of Italian populations and the geographical, ecological and cultural factors that have characterized their evolutionary history. Human presence in Italian territory is deeply rooted in the past. Lithic artifacts produced by the genus Homo and remains of Homo sapiens are among the earliest to have been found on the continent, as shown by the lithic industry of Pirro Nord (between 1.3 and 1.6 Mya) and the dental remains of the "Grotta del Cavallo" (between 45 and 43 Kya). Genetic and genomic studies relating to existing and extinct human groups have shed light on the migrations from Europe, Africa and Asia that created the ancient layers of the genetic structure of today's Italian populations, especially before the Iron Age. The important role of isolation (genetic and cultural) in shaping genetic structure is clearly visible in the patterns of intra- and inter-population diversity observed among Italian ethno-linguistic minorities that settled on the peninsula and on the major islands until the 19th century. Finally, selective pressures have likely driven the distribution of originally adaptive variants and haplotypes that now confer protection or susceptibility to major diseases such as diabetes and cardiovascular disease (in northern Italy) and tuberculosis and leprosy (in the south). What emerges is a picture where the combined effects of migration, isolation and natural selection generated by the interplay of geography, environment and culture have shaped a complex pattern of human diversity that is unique in Europe and which goes hand in hand with today's rich animal and plant biodiversity. In a nutshell, scientific evidence and cultural heritage paint Italy as a place with extremely diverse environments where distant peoples have met since the deep past, bringing and sharing genes and ideas.

Ancient genomes reveal structural shifts after the arrival of Steppe-related ancestry in the Italian Peninsula.

Across Europe, the genetics of the Chalcolithic/Bronze Age transition is increasingly characterized in terms of an influx of Steppe-related ancestry. The effect of this major shift on the genetic structure of populations in the Italian Peninsula remains underexplored. Here, genome-wide shotgun data for 22 individuals from commingled cave and single burials in Northeastern and Central Italy dated between 3200 and 1500 BCE provide the first genomic characterization of Bronze Age individuals (n = 8; 0.001-1.2× coverage) from the central Italian Peninsula, filling a gap in the literature between 1950 and 1500 BCE. Our study confirms a diversity of ancestry components during the Chalcolithic and the arrival of Steppe-related ancestry in the central Italian Peninsula as early as 1600 BCE, with this ancestry component increasing through time. We detect close patrilineal kinship in the burial patterns of Chalcolithic commingled cave burials and a shift away from this in the Bronze Age (2200-900 BCE) along with lowered runs of homozygosity, which may reflect larger changes in population structure. Finally, we find no evidence that the arrival of Steppe-related ancestry in Central Italy directly led to changes in frequency of 115 phenotypes present in the dataset, rather that the post-Roman Imperial period had a stronger influence, particularly on the frequency of variants associated with protection against Hansen's disease (leprosy). Our study provides a closer look at local dynamics of demography and phenotypic shifts as they occurred as part of a broader phenomenon of widespread admixture during the Chalcolithic/Bronze Age transition.

Insights from ancient DNA analysis of Egyptian human mummies: clues to disease and kinship.

The molecular Egyptology field started in the mid-eighties with the first publication on the ancient DNA (aDNA) analysis of an Egyptian mummy. Egypt has been a major interest for historians, archeologists, laymen as well as scientists. The aDNA research on Egyptian biological remains has been fueled by their abundance and relatively well-preserved states through artificial mummification and by the advanced analytical techniques. Early doubts of aDNA integrity within the Egyptian mummies and data authenticity were later abated with studies proving successfully authenticated aDNA retrieval. The current review tries to recapitulate the published studies presenting paleogenomic evidence of disease diagnosis and kinship establishment for the Egyptian human remains. Regarding disease diagnosis, the prevailing literature was on paleogenomic evidence of infectious diseases in the human remains. A series of reports presented evidence for the presence of tuberculosis and/or malaria. In addition, there were solitary reports of the presence of leprosy, diphtheria, bacteremia, toxoplasmosis, schistosomiasis and leishmaniasis. On the contrary, paleogenomic evidence of the presence of rare diseases was quite scarce and mentioned only in two articles. On the other hand, kinship analysis of Egyptian human remains, including that of Tutankhamen, was done using both mitochondrial DNA sequences and nuclear DNA markers, to establish family relationships in four studies. It is clear that the field of molecular Egyptology is still a largely unexplored territory. Nevertheless, the paleogenomic investigation of Egyptian remains could make significant contributions to biomedical sciences (e.g. elucidation of coevolution of human host-microbe interrelationship) as well as to evidence-based archeology.

Genetic evidence of African slavery at the beginning of the trans-Atlantic slave trade.

An archaeological excavation in Valle da Gafaria (Lagos, Portugal), revealed two contiguous burial places outside the medieval city walls, dating from the 15(th)-17(th) centuries AD: one was interpreted as a Leprosarium cemetery and the second as an urban discard deposit, where signs of violent, unceremonious burials suggested that these remains may belong to slaves captured in Africa by the Portuguese. We obtained random short autosomal sequence reads from seven individuals: two from the latter site and five from the Leprosarium and used these to call SNP identities and estimate ancestral affinities with modern reference data. The Leprosarium site samples were less preserved but gave some probability of both African and European ancestry. The two discard deposit burials each gave African affinity signals, which were further refined toward modern West African or Bantu genotyped samples. These data from distressed burials illustrate an African contribution to a low status stratum of Lagos society at a time when this port became a hub of the European trade in African slaves which formed a precursor to the transatlantic transfer of millions.

CUBN and NEBL common variants in the chromosome 10p13 linkage region are associated with multibacillary leprosy in Vietnam.

Leprosy is caused by infection with Mycobacterium leprae and is classified clinically into paucibacillary (PB) or multibacillary (MB) subtypes based on the number of skin lesions and the bacillary index detected in skin smears. We previously identified a major PB susceptibility locus on chromosome region 10p13 in Vietnamese families by linkage analysis. In the current study, we conducted high-density association mapping of the 9.5 Mb linkage peak on chromosome region 10p13 covering 39 genes. Using leprosy per se and leprosy subtypes as phenotypes, we employed 294 nuclear families (303 leprosy cases, 63 % MB, 37 % PB) as a discovery sample and 192 nuclear families (192 cases, 55 % MB, 45 % PB) as a replication sample. Replicated significant association signals were revealed in the genes for cubilin (CUBN) and nebulette (NEBL). In the combined sample, the C allele (frequency 0.26) at CUBN SNP rs10904831 showed association [p = 1 × 10(-5); OR 0.52 (0.38-0.7)] with MB leprosy only. Likewise, allele T (frequency 0.42) at NEBL SNP rs11012461 showed association [p = 4.2 × 10(-5); OR 2.51 (1.6-4)] with MB leprosy only. These associations remained valid for the CUBN signal when taking into account the effective number of tests performed (type I error significance threshold = 2.4 × 10(-5)). We used the results of our analyses to propose a new model for the genetic control of polarization of clinical leprosy.

Palaeopathology and genes: investigating the genetics of infectious diseases in excavated human skeletal remains and mummies from past populations.

The aim of this paper is to review the use of genetics in palaeomicrobiology, and to highlight the importance of understanding past diseases. Palaeomicrobiology is the study of disease pathogens in skeletal and mummified remains from archaeological contexts. It has revolutionarised our understanding of health in the past by enabling a deeper knowledge of the origins and evolution of many diseases that have shaped us as a species. Bacterial diseases explored include tuberculosis, leprosy, bubonic plague, typhoid, syphilis, endemic and epidemic typhus, trench fever, and Helicobacter pylori. Viral diseases discussed include influenza, hepatitis B, human papilloma virus (HPV), human T-cell lymphotrophic virus (HTLV-1) and human immunodeficiency virus (HIV). Parasitic diseases investigated include malaria, leishmaniasis, Chagas' disease, roundworm, whipworm, pinworm, Chinese liver fluke, fleas and lice. Through a better understanding of disease origins and their evolution, we can place into context how many infectious diseases are changing over time, and so help us estimate how they may change in the future.

Tracking climate change in a dispersal-limited species: reduced spatial and genetic connectivity in a montane salamander.

Tropical montane taxa are often locally adapted to very specific climatic conditions, contributing to their lower dispersal potential across complex landscapes. Climate and landscape features in montane regions affect population genetic structure in predictable ways, yet few empirical studies quantify the effects of both factors in shaping genetic structure of montane-adapted taxa. Here, we considered temporal and spatial variability in climate to explain contemporary genetic differentiation between populations of the montane salamander, Pseudoeurycea leprosa. Specifically, we used ecological niche modelling (ENM) and measured spatial connectivity and gene flow (using both mtDNA and microsatellite markers) across extant populations of P. leprosa in the Trans-Mexican Volcanic Belt (TVB). Our results indicate significant spatial and genetic isolation among populations, but we cannot distinguish between isolation by distance over time or current landscape barriers as mechanisms shaping population genetic divergences. Combining ecological niche modelling, spatial connectivity analyses, and historical and contemporary genetic signatures from different classes of genetic markers allows for inference of historical evolutionary processes and predictions of the impacts future climate change will have on the genetic diversity of montane taxa with low dispersal rates. Pseudoeurycea leprosa is one montane species among many endemic to this region and thus is a case study for the continued persistence of spatially and genetically isolated populations in the highly biodiverse TVB of central Mexico.

Nuclear and chloroplast DNA phylogeography reveals Pleistocene divergence and subsequent secondary contact of two genetic lineages of the tropical rainforest tree species Shorea leprosula (Dipterocarpaceae) in South-East Asia.

Tropical rainforests in South-East Asia have been affected by climatic fluctuations during past glacial eras. To examine how the accompanying changes in land areas and temperature have affected the genetic properties of rainforest trees in the region, we investigated the phylogeographic patterns of a widespread dipterocarp species, Shorea leprosula. Two types of DNA markers were used: expressed sequence tag-based simple sequence repeats and chloroplast DNA (cpDNA) sequence variations. Both sets of markers revealed clear genetic differentiation between populations in Borneo and those in the Malay Peninsula and Sumatra (Malay/Sumatra). However, in the south-western part of Borneo, genetic admixture of the lineages was observed in the two marker types. Coalescent simulation based on cpDNA sequence variation suggested that the two lineages arose 0.28-0.09 million years before present and that following their divergence migration from Malay/Sumatra to Borneo strongly exceeded migration in the opposite direction. We conclude that the genetic structure of S. leprosula was largely formed during the middle Pleistocene and was subsequently modified by eastward migration across the subaerially exposed Sunda Shelf.

Insights from genomic comparisons of genetically monomorphic bacterial pathogens.

Some of the most deadly bacterial diseases, including leprosy, anthrax and plague, are caused by bacterial lineages with extremely low levels of genetic diversity, the so-called 'genetically monomorphic bacteria'. It has only become possible to analyse the population genetics of such bacteria since the recent advent of high-throughput comparative genomics. The genomes of genetically monomorphic lineages contain very few polymorphic sites, which often reflect unambiguous clonal genealogies. Some genetically monomorphic lineages have evolved in the last decades, e.g. antibiotic-resistant Staphylococcus aureus, whereas others have evolved over several millennia, e.g. the cause of plague, Yersinia pestis. Based on recent results, it is now possible to reconstruct the sources and the history of pandemic waves of plague by a combined analysis of phylogeographic signals in Y. pestis plus polymorphisms found in ancient DNA. Different from historical accounts based exclusively on human disease, Y. pestis evolved in China, or the vicinity, and has spread globally on multiple occasions. These routes of transmission can be reconstructed from the genealogy, most precisely for the most recent pandemic that was spread from Hong Kong in multiple independent waves in 1894.

Tales from scales: old DNA yields insights into contemporary evolutionary processes affecting fishes.

Salmon and trout populations are suffering declines in abundance and diversity over much of their range around the Atlantic and Pacific rims as a consequence of many factors. One method of dealing with the decline has been to produce them in hatcheries but the wisdom of this approach has been hotly debated (e.g. Hilborn & Winton 1993; Waples 1999; Brannon et al. 2004). One concern is that domesticated hatchery strains will interbreed with locally adapted wild fish; but how do we study the genetic effects if the introgression might have occurred in the past? Hansen (2002) used DNA isolated from archived scales from brown trout, Salmo trutta (Fig. 1), to show that domesticated trout had, to varying degrees, genetically introgressed with wild, native trout in two Danish rivers. Extending that study, Hansen et al. (2009) have examined DNA from brown trout scales in six Danish rivers collected during historical (1927-1956) and contemporary (2000-2006) periods and from two hatchery source populations, to assess the effects of stocking nonlocal strains of hatchery trout and declining abundance on genetic diversity. Using 21 microsatellite loci, they revealed that genetic change occurred between the historic and contemporary time periods. Many populations appeared to have some low level of introgression from hatchery stocks and two populations apparently experienced high levels of introgression. Hansen et al. (2009) also showed that population structure persists in contemporary populations despite apparent admixture and migration among populations, providing evidence that the locally adapted populations have struggled against and, to some extent, resisted being overwhelmed by repeated introductions of and interbreeding with non-native, hatchery-produced conspecifics.

A note on the extended Rosenbrock function.

The Rosenbrock function is a well-known benchmark for numerical optimization problems, which is frequently used to assess the performance of Evolutionary Algorithms. The classical Rosenbrock function, which is a two-dimensional unimodal function, has been extended to higher dimensions in recent years. Many researchers take the high-dimensional Rosenbrock function as a unimodal function by instinct. In 2001 and 2002, Hansen and Deb found that the Rosenbrock function is not a unimodal function for higher dimensions although no theoretical analysis was provided. This paper shows that the n-dimensional (n = 4 approximately 30) Rosenbrock function has 2 minima, and analysis is proposed to verify this. The local minima in some cases are presented. In addition, this paper demonstrates that one of the "local minima" for the 20-variable Rosenbrock function found by Deb might not in fact be a local minimum.

Spatial structure and genetic diversity of two tropical tree species with contrasting breeding systems and different ploidy levels.

Analyses of the spatial distribution pattern, spatial genetic structure and of genetic diversity were carried out in two tropical tree species with contrasting breeding systems and different ploidy levels using a 50-ha demographic plot in a lowland dipterocarp forest in Peninsular Malaysia. Shorea leprosula is a diploid and predominantly outcrossed species, whereas S. ovalis ssp. sericea is an autotetraploid species with apomictic mode of reproduction. Genetic diversity parameters estimated for S. leprosula using microsatellite were consistently higher than using allozyme. In comparisons with S. leprosula and other tropical tree species, S. ovalis ssp. sericea also displayed relatively high levels of genetic diversity. This might be explained by the lower pressure of genetic drift due to tetrasomic inheritance, and for autotetraploids each locus can accommodate up to four different alleles and this allows maintenance of more alleles at individual loci. The observed high levels of genetic diversity in S. ovalis ssp. sericea can also be due to a random retention of more heterogeneous individuals in the past, and the apomictic mode of reproduction might be an evolutionary strategy, which allows the species to maintain high levels of genetic diversity. The spatial distribution pattern analyses of both species showed significant levels of aggregation at small and medium but random distribution at the big diameter-class. The decrease in magnitude of spatial aggregation from small- to large-diameter classes might be due to compensatory mortality during recruitment and survival under competitive thinning process. Spatial genetic structure analyses for both species revealed significant spatial genetic structure for short distances in all the three diameter-classes. The magnitude of spatial genetic structure in both species was observed to be decreasing from smaller- to larger-diameter classes. The high spatial genetic structuring observed in S. ovalis ssp. sericea at the small-diameter class is due primarily to limited seed dispersal and apomictic mode of reproduction. The similar observation in S. leprosula, however, can be explained by limited seed and pollen dispersal, which supports further the fact that the species is pollinated by weak fliers, mainly of Thrips and Megalurothrips in the lowland dipterocarp forest.

Evidence for a cluster of genes on chromosome 17q11-q21 controlling susceptibility to tuberculosis and leprosy in Brazilians

The region of conserved synteny on mouse chromosome 11/human 17q11-q21 is known to carry a susceptibility gene(s) for intramacrophage pathogens. The region is rich in candidates including NOS2A, CCL2/MCP-1, CCL3/MIP-1 alpha, CCL4/MIP-1 beta, CCL5/RANTES, CCR7, STAT5A/5B. To examine the region in man, we studied 92 multicase tuberculosis (627 individuals) and 72 multicase leprosy (372 indiciduals) families from Brazil. Multipoint nonparametric analysis (ALLEGRO) using 16 microsatellites shows two peaks of linkage for leprosy at D17S250 (Zir score 2.34; P=0.01) and D17S1795 (Zir 2.67; P=O.004) and a single peack for tuberculosis at D17S250 (Zir 2.04; P=0.02). Combined analysis shows significant linkage (peak Zir 3.38) at D17S250, equivalent to an allele sharing LOD score 2.48 (P=0.0004). To determine whether one or multiple genes contribute, 49 informative single nucleotide polymorphisms were typed in candidate genes. Family-based allelic association testing that was robust to family clustering demonstrated significant associations with tuberculosis susceptibility at four loci separated by intervals (NOS2A-8.4 Mb-CCL 18-32.3 kb-CCL4-6.04 Mb-STAT5B) up to several Mb. Stepwise conditional logistic regression analysis using a case/pseudo-control data set showed that the four genes contributed separate main effects, consistent with a cluster of susceptibilitty genes acros 17q11.2

A region of chromosome 20 is linked to leprosy susceptibility in a South Indian population.

A major susceptibility locus for leprosy has recently been mapped on chromosome 10 (10p13) by genome-wide linkage analysis. Microsatellite markers from this genome screen that showed suggestive evidence of linkage to leprosy were evaluated in an additional 140 families with affected sib pairs. A second region of linkage has thus been identified on chromosome 20 (20p12). The peak of linkage lies at marker D20S115, which has a significant single-point maximum logarithm of odds score of 3.48 (P=.00003). Transmission disequilibrium testing of the microsatellite markers in 20p12 showed that the marker D20S835 is associated with protection against leprosy (P=.021), which suggests that a locus controlling susceptibility lies close to this marker.

Rheumatic disorders in Sub-saharan Africa

OBJECTIVE: To review prevalence of rheumatic disorders in Sub-saharan Africa and in the context of current medical practice in the region assess the need for service and educational provision. DATA SOURCES: Medline, (English, French). Pre-Medline literature review from the 1950's (Current contents). Various conference reports including attendance at all three AFLAR (African League Against Rheumatism) congresses in the 1990's. Author's personal database. All cited references read in full. CONCLUSIONS: The evidence shows rheumatoid arthritis and systemic lupus erythematosus to be increasing in frequency in the indigenous populations of East, Central and South Africa but remaining rare in West Africans. Gout is now more prevalent than ever throughout the subcontinent. HIV has spawned a variety of previously rare spondyloarthropathies (reactive arthritis, psoriatic arthritis, enthesopathy) and changed the epidemiology of pyomyositis and osteomyelitis. Osteoarthritis is a universal problem. Juvenile chronic arthritis is not rare and rheumatic fever is common. Acute and chronic locomotor problems associated with diverse entities such as leprosy, brucellosis, meningococcus, alpha viruses, parasites, fluorosis, rickets and haemoglobinopathies enhance diagnostic diversity and therapeutic and educational requirements. Suggestions made to address the challenge posed by the burden of rheumatic disorders.

Rheumatic disorders in Sub-saharan Africa.

OBJECTIVE: To review prevalence of rheumatic disorders in Sub-saharan Africa and in the context of current medical practice in the region assess the need for service and educational provision. DATA SOURCES: Medline, (English, French). Pre-Medline literature review from the 1950's (Current contents). Various conference reports including attendance at all three AFLAR (African League Against Rheumatism) congresses in the 1990's. Author's personal database. All cited references read in full. CONCLUSIONS: The evidence shows rheumatoid arthritis and systemic lupus erythematosus to be increasing in frequency in the indigenous populations of East, Central and South Africa but remaining rare in West Africans. Gout is now more prevalent than ever throughout the subcontinent. HIV has spawned a variety of previously rare spondyloarthropathies (reactive arthritis, psoriatic arthritis, enthesopathy) and changed the epidemiology of pyomyositis and osteomyelitis. Osteoarthritis is a universal problem. Juvenile chronic arthritis is not rare and rheumatic fever is common. Acute and chronic locomotor problems associated with diverse entities such as leprosy, brucellosis, meningococcus, alpha viruses, parasites, fluorosis, rickets and haemoglobinopathies enhance diagnostic diversity and therapeutic and educational requirements. Suggestions made to address the challenge posed by the burden of rheumatic disorders.

A region of chromosome 20 is linked to leprosy susceptibility in a South Indian population

A major susceptibility locus for leprosy has recently been mapped on chromosome 10 (10p13) by genome-wide linkage analysis. Microsatellite markers from this genome screen that showed suggestive evidence of linkage to leprosy were evaluated in an additional 140 families with affected sib pairs. A second region of linkage has thus been identified on chromosome 20 (20p12). The peak of linkage lies at marker D20S115, which has a significant single-point maximum logarithm of odds score of 3.48 (P=.00003). Transmission disequilibrium testing of the microsatellite markers in 20p12 showed that the marker D20S835 is associated with protection against leprosy (P=.021), which suggests that a locus controlling susceptibility lies close to this marker.

Paternity testing with VNTR DNA systems. II. Evaluation of 271 cases of disputed paternity with the VNTR systems D2S44, D5S43, D7S21, D7S22, and D12S11.

Paternity testing was carried out in 271 cases of disputed paternity using the 5 VNTR systems D2S44 (YNH24), D5S43 (MS8), D7S21 (MS31), D7S22 (g3), and D12S11 (MS43a), and 10-15 conventional marker systems including the HLA-A,B system. By means of the matching criteria for the VNTR systems established elsewhere (Morling & Hansen 1992), all 70 unrelated men who had been excluded by conventional typing were also excluded with 2 or more VNTR systems. Based on the observed exclusion frequencies for the 5 VNTR systems, a theoretical exclusion rate exceeding 0.999 could be obtained. A total of 350 father/child pairs were studied and in 3 paternity cases and one immigrant family, the alleged fathers were excluded solely by one of the 5 VNTR systems possibly reflecting mutations. No mother/child exclusions were observed among 350 mother/child pairs. Linkage analysis between the syntenic systems D7S21 (MS31) and D7S22 (g3) was performed in 29 informative families with 81 children and revealed a recombination distance of about 31 cM. The positive evidence for paternity provided by the 5 VNTR systems in cases with non-exclusions is discussed.

HLA-DR-associated genetic control of the type of leprosy in a population from surinam.

The relationship between HLA phenotype and leprosy classification was studied in 73 unrelated patients and 92 healthy controls from a mixed Negroid-Caucasoid population originating from Surinam, South America. Heterogeneity in the distribution of HLA-DR (but not A, B, and C) was detected between tuberculoid (TT* + BT*) leprosy and lepromatous (BL* + LL*) leprosy patients (p = 0.024). This heterogeneity appeared to be caused almost exclusively by DR3. Most significantly, the frequency of DR3 was increased among polar tuberculoid (TT) leprosy patients as compared to the rest of the patients (p = 0.0003). Compared with healthy controls the frequency of DR3 was increased among TT patients (p = 0.006), unchanged in BT patients, and decreased among lepromatous (BL + LL) patients (p = 0.027). These data indicate that in this population an DR3-associated factor controls the type of the disease that develops after infection with Mycobacterium leprae.