The saliva microbiome of Pan and Homo.

BACKGROUND: It is increasingly recognized that the bacteria that live in and on the human body (the microbiome) can play an important role in health and disease. The composition of the microbiome is potentially influenced by both internal factors (such as phylogeny and host physiology) and external factors (such as diet and local environment), and interspecific comparisons can aid in understanding the importance of these factors. RESULTS: To gain insights into the relative importance of these factors on saliva microbiome diversity, we here analyze the saliva microbiomes of chimpanzees (Pan troglodytes) and bonobos (Pan paniscus) from two sanctuaries in Africa, and from human workers at each sanctuary. The saliva microbiomes of the two Pan species are more similar to one another, and the saliva microbiomes of the two human groups are more similar to one another, than are the saliva microbiomes of human workers and apes from the same sanctuary. We also looked for the existence of a core microbiome and find no evidence for a taxon-based core saliva microbiome for Homo or Pan. In addition, we studied the saliva microbiome from apes from the Leipzig Zoo, and found an extraordinary diversity in the zoo ape saliva microbiomes that is not found in the saliva microbiomes of the sanctuary animals. CONCLUSIONS: The greater similarity of the saliva microbiomes of the two Pan species to one another, and of the two human groups to one another, are in accordance with both the phylogenetic relationships of the hosts as well as with host physiology. Moreover, the results from the zoo animals suggest that novel environments can have a large impact on the microbiome, and that microbiome analyses based on captive animals should be viewed with caution as they may not reflect the microbiome of animals in the wild.

High diversity of the saliva microbiome in Batwa Pygmies.

We describe the saliva microbiome diversity in Batwa Pygmies, a former hunter-gatherer group from Uganda, using next-generation sequencing of partial 16S rRNA sequences. Microbial community diversity in the Batwa is significantly higher than in agricultural groups from Sierra Leone and the Democratic Republic of Congo. We found 40 microbial genera in the Batwa, which have previously not been described in the human oral cavity. The distinctive composition of the salvia microbiome of the Batwa may have been influenced by their recent different lifestyle and diet.

High-throughput sequencing of complete human mtDNA genomes from the Caucasus and West Asia: high diversity and demographic inferences.

To investigate the demographic history of human populations from the Caucasus and surrounding regions, we used high-throughput sequencing to generate 147 complete mtDNA genome sequences from random samples of individuals from three groups from the Caucasus (Armenians, Azeri and Georgians), and one group each from Iran and Turkey. Overall diversity is very high, with 144 different sequences that fall into 97 different haplogroups found among the 147 individuals. Bayesian skyline plots (BSPs) of population size change through time show a population expansion around 40-50 kya, followed by a constant population size, and then another expansion around 15-18 kya for the groups from the Caucasus and Iran. The BSP for Turkey differs the most from the others, with an increase from 35 to 50 kya followed by a prolonged period of constant population size, and no indication of a second period of growth. An approximate Bayesian computation approach was used to estimate divergence times between each pair of populations; the oldest divergence times were between Turkey and the other four groups from the South Caucasus and Iran (~400-600 generations), while the divergence time of the three Caucasus groups from each other was comparable to their divergence time from Iran (average of ~360 generations). These results illustrate the value of random sampling of complete mtDNA genome sequences that can be obtained with high-throughput sequencing platforms.

A Y-STR database of Iranian and Azerbaijanian minority populations.

Seventeen Y-chromosomal short tandem repeats (Y-STR) DYS19, DYS389I, DYS389II, DYS390, DYS391, DYS392, DYS393, DYS385a, DYS385b, DYS437, DYS438, DYS439, DYS448, DYS456, DYS458, DYS635 and GATA H4 were studied in five minor linguistic groups from Iran (Arabs, Gilaki, Mazandarani, Bakhtiari and Southern Talysh) and one from Azerbaijan (Northern Talysh) with the goal of constructing of a representative Y-STR database for this region in Southwest Asia. Analysis of Molecular Variance (AMOVA) reveals non-significant or low genetic distances between the Iranian Gilaki, Mazandarani, Bakhtiari and non-Iranian Turkish, Azerbaijanian, Armenian and Kurd populations, but larger genetic distances to both Talysh populations, the Iranian Arabs, Georgian and Kazakh populations.

Comparative analysis of human saliva microbiome diversity by barcoded pyrosequencing and cloning approaches.

Metagenomic studies traditionally rely on cloning polymerase chain reaction (PCR) products and sequencing multiple clones. However, this approach is tedious and expensive, thereby limiting the range and scale of questions that can be addressed. Recent developments in DNA sequencing technologies enable a dramatic increase in throughput via parallel in-depth analysis of many samples with limited sample processing and lower costs. We directly compared the traditional cloning approach with a barcoded pyrosequencing method to see whether the latter accurately describes microbiome diversity in human saliva. Our results indicate that despite the shorter read lengths, the pyrosequencing approach provides a description of the human salivary microbiome that is in good agreement with results based on the traditional cloning and sequencing approach.

Global diversity in the human salivary microbiome.

The human salivary microbiome may play a role in diseases of the oral cavity and interact with microbiomes from other parts of the human body (in particular, the intestinal tract), but little is known about normal variation in the salivary microbiome. We analyzed 14,115 partial ( approximately 500 bp) 16S ribosomal RNA (rRNA) sequences from saliva samples from 120 healthy individuals (10 individuals from each of 12 worldwide locations). These sequences could be assigned to 101 known bacterial genera, of which 39 were not previously reported from the human oral cavity; phylogenetic analysis suggests that an additional 64 unknown genera are present. There is high diversity in the salivary microbiome within and between individuals, but little geographic structure. Overall, approximately 13.5% of the total variance in the composition of genera is due to differences among individuals, which is remarkably similar to the fraction of the total variance in neutral genetic markers that can be attributed to differences among human populations. Investigation of some environmental variables revealed a significant association between the genetic distances among locations and the distance of each location from the equator. Further characterization of the enormous diversity revealed here in the human salivary microbiome will aid in elucidating the role it plays in human health and disease, and in the identification of potentially informative species for studies of human population history.

Genomic complexity of the Y-STR DYS19: inversions, deletions and founder lineages carrying duplications.

The Y-STR DYS19 is firmly established in the repertoire of Y-chromosomal markers used in forensic analysis yet is poorly understood at the molecular level, lying in a complex genomic environment and exhibiting null alleles, as well as duplications and occasional triplications in population samples. Here, we analyse three null alleles and 51 duplications and show that DYS19 can also be involved in inversion events, so that even its location within the short arm of the Y chromosome is uncertain. Deletion mapping in the three chromosomes carrying null alleles shows that their deletions are less than approximately 300 kb in size. Haplotypic analysis with binary markers shows that they belong to three different haplogroups and so represent independent events. In contrast, a collection of 51 DYS19 duplication chromosomes belong to only four haplogroups: two are singletons and may represent somatic mutation in lymphoblastoid cell lines, but two, in haplogroups G and C3c, represent founder lineages that have spread widely in Central Europe/West Asia and East Asia, respectively. Consideration of candidate mechanisms underlying both deletions and duplications provides no evidence for the involvement of non-allelic homologous recombination, and they are likely to represent sporadic events with low mutation rates. Understanding the basis and population distribution of these DYS19 alleles will aid in the utilisation and interpretation of profiles that contain them.

mtDNA and Y-chromosome variation in the Talysh of Iran and Azerbaijan.

The Northern Talysh from Azerbaijan and the Southern Talysh from Iran self-identify as one ethnic group and speak a Northwestern Iranian language. However, the Northern and Southern Talysh dialects are so different that they may actually be separate languages. Does this linguistic differentiation reflect internal change due to isolation, or could contact-induced change have played a role? We analyzed mtDNA HVI sequences, 11 Y-chromosome bi-allelic markers, and 9 Y-STR loci in Northern and Southern Talysh and compared them with their neighboring groups. The mtDNA data show a close relatedness of both groups with each other and with neighboring groups, whereas the Northern Talysh Y-chromosome variation differs from that of neighboring groups, probably as a result of genetic drift. This genetic drift most likely reflects a founder event in the male gene pool of Northern Talysh: either fewer males than females migrated to Azerbaijan, or there was a higher degree of relatedness among the male migrants. Since we find no evidence of substantial genetic contact between either Northern or Southern Talysh and neighboring groups, we conclude that internal change, rather than contact-induced change, most likely explains the linguistic differentiation between Northern and Southern Talysh.

Boundaries and clines in the West Eurasian Y-chromosome landscape: insights from the European part of Russia.

Previous studies of Y chromosome variation have revealed that western Europe, the Volga-Ural region, and the Caucasus differ dramatically with respect to Y-SNP haplogroup composition. The European part of Russia is situated in between these three regions; to determine if these differences reflect clines or boundaries in the Y-chromosome landscape, we analyzed 12 Y-SNPs in 545 males from 12 populations from the European part of Russia. The majority of Russian Y chromosomes (from 74% to 94%) belong to three Y chromosomal lineages [I-M170, R1a1-M17, and N3-TAT] that are also frequent in the rest of east Europe, north Europe, and/or in the Volga-Ural region. We find significant but low correlations between haplogroup frequencies and the geographic location of populations, suggesting gradual change in the Y chromosome gene pool across western Eurasia. However, we also find some significant boundaries between populations, suggesting that both isolation and migration have influenced the Y chromosome landscape.

Dynamic nature of the proximal AZFc region of the human Y chromosome: multiple independent deletion and duplication events revealed by microsatellite analysis.

The human Y chromosome shows frequent structural variants, some of which are selectively neutral, while others cause impaired fertility due to the loss of spermatogenic genes. The large-scale use of multiple Y-chromosomal microsatellites in forensic and population genetic studies can reveal such variants, through the absence or duplication of specific markers in haplotypes. We describe Y chromosomes in apparently normal males carrying null and duplicated alleles at the microsatellite DYS448, which lies in the proximal part of the azoospermia factor c (AZFc) region, important in spermatogenesis, and made up of "ampliconic" repeats that act as substrates for nonallelic homologous recombination (NAHR). Physical mapping in 26 DYS448 deletion chromosomes reveals that only three cases belong to a previously described class, representing independent occurrences of an approximately 1.5-Mb deletion mediated by recombination between the b1 and b3 repeat units. The remainder belong to five novel classes; none appears to be mediated through homologous recombination, and all remove some genes, but are likely to be compatible with normal fertility. A combination of deletion analysis with binary-marker and microsatellite haplotyping shows that the 26 deletions represent nine independent events. Nine DYS448 duplication chromosomes can be explained by four independent events. Some lineages have risen to high frequency in particular populations, in particular a deletion within haplogroup (hg) C(*)(xC3a,C3c) found in 18 Asian males. The nonrandom phylogenetic distribution of duplication and deletion events suggests possible structural predisposition to such mutations in hgs C and G.

Analysis of Y chromosome STR haplotypes in the European part of Russia reveals high diversities but non-significant genetic distances between populations.

A total of 17 Y-specific STR loci were studied in 12 districts of the European part of Russia aiming to ascertain the amount of substructure required for the construction of a representative regional database. All groups exhibited high haplotype diversities but low inter-population variance as measured by an analysis of molecular variance. However, when Western Russia is taken as a whole, the genetic distances to the neighbouring populations were significant. Whereas gradual change in the Y chromosome pool exists between Russia and the Slavic-speaking populations to the West, remarkable discontinuities were observed with neighbouring populations in the East, North and South.

Y-chromosomal STR haplotypes in Kalmyk population samples.

Seventeen Y-chromosomal short tandem repeats (STRs), DYS19, DYS389I, DYS389II, DYS390, DYS391, DYS392, DYS393, DYS385ab, DYS437, DYS438, DYS439, GATA-H4, DYS448, DYS456, DYS458, DYS635 were typed in DNA samples from the Kalmyk population (n=99). The population is characterized by a high proportion of duplicated DYS19 alleles and deletions of the locus DYS448 on the background of the Central Asian haplogroup C*. AMOVA analysis reveals a close vicinity to Mongolian and Kazakh populations and large genetic distance to geographical neighbours from Russia, Ukraine and the Caucasus.

The Gagauz, a linguistic enclave, are not a genetic isolate.

The Gagauz are a Turkic-speaking group that migrated from Turkey to their present location in the southern part of the Republic of Moldova about 150 years ago. Surrounded by Indo-European-speaking populations, they thus form a linguistic enclave, which raises the following question: to what extent have they remained in genetic isolation from their geographic neighbours? Analyses of mtDNA and Y chromosome variation indicate that despite their linguistic differences, the Gagauz have admixed extensively with neighbouring groups. Our data suggest that there has been more mtDNA than Y chromosome admixture, in keeping with the patrilocal nature of these groups. Moreover, when compared with another linguistic enclave, the Kalmyks there appears to be a correlation between the amount of genetic admixture and the amount of linguistic influence that these two linguistic enclaves have experienced from neighbouring groups.

Concomitant replacement of language and mtDNA in South Caspian populations of Iran.

The Gilaki and Mazandarani occupy the South Caspian region of Iran and speak languages belonging to the North-Western branch of Iranian languages . It has been suggested that their ancestors came from the Caucasus region, perhaps displacing an earlier group in the South Caspian . Linguistic evidence supports this scenario, in that the Gilaki and Mazandarani languages (but not other Iranian languages) share certain typological features with Caucasian languages . We analyzed patterns of mtDNA and Y chromosome variation in the Gilaki and Mazandarani. Based on mtDNA HV1 sequences, the Gilaki and Mazandarani most closely resemble their geographic and linguistic neighbors, namely other Iranian groups. However, their Y chromosome types most closely resemble those found in groups from the South Caucasus. A scenario that explains these differences is a south Caucasian origin for the ancestors of the Gilaki and Mazandarani, followed by introgression of women (but not men) from local Iranian groups, possibly because of patrilocality. Given that both mtDNA and language are maternally transmitted, the incorporation of local Iranian women would have resulted in the concomitant replacement of the ancestral Caucasian language and mtDNA types of the Gilaki and Mazandarani with their current Iranian language and mtDNA types. Concomitant replacement of language and mtDNA may be a more general phenomenon than previously recognized.

Evaluation of saliva as a source of human DNA for population and association studies.

A simple noninvasive procedure for saliva sample collection and DNA extraction was developed. On average, the amount of human DNA (as measured by a TaqMan-based assay) was about 11.4 microg/mL saliva, which is more than can be obtained from other noninvasive samples such as cheek swabs. However, the presence of large amounts of nonhuman DNA (up to 90% of the total extracted DNA) in saliva samples does necessitate DNA quantitation methods that are specific for human DNA. We were able to reliably and accurately type different genetic markers (mDNA sequences, Y-chromosomal single-nucleotide polymorphisms, and autosomal microsatellite loci) from saliva samples stored for up to 30 days at 37 degrees C, making this method well-suited for field conditions and convenient transportation of samples back to the laboratory. Thus, saliva can be considered a reliable source of DNA for a wide variety of genetic studies.

Genetic evidence for the Mongolian ancestry of Kalmyks.

The Kalmyks are an ethnic group along the lower Volga River in Russia who are thought to have migrated there from Mongolia about 300 years ago. To investigate their origins, we studied mtDNA and Y-chromosome variation in 99 Kalmyks. Both mtDNA HV1 sequences and Y-chromosome SNP haplogroups indicate a close relationship of Kalmyks with Mongolians. In addition, genetic diversity for both mtDNA and the Y chromosome are comparable in Kalmyks, Mongolians, and other Central Asian groups, indicating that the Kalmyk migration was not associated with a substantial bottleneck. The so-called "Genghis Khan" Y-chromosome short tandem repeat (STR) haplotype was found in high frequency (31.3%) among Kalmyks, further supporting a strong genetic connection between Kalmyks and Mongolians. Genetic analyses of even recent, relatively well-documented migrations such as of the Kalmyks can therefore lead to new insights concerning such migrations.

MtDNA and Y-chromosome variation in Kurdish groups.

In order to investigate the origins and relationships of Kurdish-speaking groups, mtDNA HV1 sequences, eleven Y chromosome bi-allelic markers, and 9 Y-STR loci were analyzed among three Kurdish groups: Zazaki and Kurmanji speakers from Turkey, and Kurmanji speakers from Georgia. When compared with published data from other Kurdish groups and from European, Caucasian, and West and Central Asian groups, Kurdish groups are most similar genetically to other West Asian groups, and most distant from Central Asian groups, for both mtDNA and the Y-chromosome. However, Kurdish groups show a closer relationship with European groups than with Caucasian groups based on mtDNA, but the opposite based on the Y-chromosome, indicating some differences in their maternal and paternal histories. The genetic data indicate that the Georgian Kurdish group experienced a bottleneck effect during their migration to the Caucasus, and that they have not had detectable admixture with their geographic neighbours in Georgia. Our results also do not support the hypothesis of the origin of the Zazaki-speaking group being in northern Iran; genetically they are more similar to other Kurdish groups. Genetic analyses of recent events, such as the origins and migrations of Kurdish-speaking groups, can therefore lead to new insights into such migrations.

Haplotypes from the Caucasus, Turkey and Iran for nine Y-STR loci.

Nine Y chromosome short tandem repeat (STR) loci (DYS385a, DYS385b, DYS389I, DYS389II, DYS390, DYS391, DYS392, DYS393 and DYS19 (DYS394)) were typed in 669 individuals belonging to 16 populations from the Caucasus, Turkey and Iran.

Testing hypotheses of language replacement in the Caucasus: evidence from the Y-chromosome.

A previous analysis of mtDNA variation in the Caucasus found that Indo-European-speaking Armenians and Turkic-speaking Azerbaijanians were more closely related genetically to other Caucasus populations (who speak Caucasian languages) than to other Indo-European or Turkic groups, respectively. Armenian and Azerbaijanian therefore represent language replacements, possibly via elite dominance involving primarily male migrants, in which case genetic relationships of Armenians and Azerbaijanians based on the Y-chromosome should more closely reflect their linguistic relationships. We therefore analyzed 11 bi-allelic Y-chromosome markers in 389 males from eight populations, representing all major linguistic groups in the Caucasus. As with the mtDNA study, based on the Y-chromosome Armenians and Azerbaijanians are more closely-related genetically to their geographic neighbors in the Caucasus than to their linguistic neighbors elsewhere. However, whereas the mtDNA results show that Caucasian groups are more closely related genetically to European than to Near Eastern groups, by contrast the Y-chromosome shows a closer genetic relationship with the Near East than with Europe.