Characterization of the complete mitochondrial genome and phylogenetic analyses of Haemaphysalis tibetensis Hoogstraal, 1965 (Acari: Ixodidae).

Ticks are specialized ectoparasites that feed on blood, causing physical harm to the host and facilitating pathogen transmission. The genus Haemaphysalis contains vectors for numerous infectious agents. These agents cause various diseases in humans and animals. Mitochondrial genome sequences serve as reliable molecular markers, forming a crucial basis for evolutionary analyses, studying species origins, and exploring molecular phylogeny. We extracted mitochondrial genome from the enriched mitochondria of Haemaphysalis tibetensis and obtained a 14,714-bp sequence. The mitochondrial genome consists of 13 protein-coding genes (PCGs), two ribosomal RNA, 22 transfer RNAs (tRNAs), and two control regions. The nucleotide composition of H. tibetensis mitochondrial genome was 38.38 % for A, 9.61 % for G, 39.32 % for T, and 12.69 % for C. The A + T content of H. tibetensis mitochondrial genome was 77.7 %, significantly higher than the G + C content. The repeat units of H. tibetensis exhibited two identical repeat units of 33 bp in length, positioned downstream of nad1 and rrnL genes. Furthermore, phylogenetic analyses based on the 13 PCGs indicated that Haemaphysalis tibetensis (subgenus Allophysalis) formed a monophyletic clade with Haemaphysalis nepalensis (subgenus Herpetobia) and Haemaphysalis danieli (subgenus Allophysalis). Although the species Haemaphysalis inermis, Haemaphysalis kitaokai, Haemaphysalis kolonini, and Haemaphysalis colasbelcouri belong to the subgenus Alloceraea, which were morphologically primitive hemaphysalines just like H. tibetensis, these four tick species cannot form a single clade with H. tibetensis. In this study, the whole mitochondrial genome sequence of H. tibetensis from Tibet was obtained, which enriched the mitochondrial genome data of ticks and provided genetic markers to study the population heredity and molecular evolution of the genus Haemaphysalis.

The complete mitogenome of the Asian lung fluke Paragonimus skrjabini miyazakii and its implications for the family Paragonimidae (Trematoda: Platyhelminthes).

The complete circular mitogenome of Paragonimus skrjabini miyazakii (Platyhelminthes: Paragonimidae) from Japan, obtained by PacBio long-read sequencing, was 17 591 bp and contained 12 protein-coding genes (PCGs), 2 mitoribosomal RNA and 22 transfer RNA genes. The atp8 gene was absent, and there was a 40 bp overlap between nad4L and nad4. The long non-coding region (4.3 kb) included distinct types of long and short repeat units. The pattern of base usage for PCGs and the mtDNA coding region overall in Asian and American Paragonimus species (P. s. miyazakii, P. heterotremus, P. ohirai and P. kellicotti) and the Indian form of P. westermani was T > G > A > C. On the other hand, East-Asian P. westermani used T > G > C > A. Five Asian and American Paragonimus species and P. westermani had TTT/Phe, TTG/Leu and GTT/Val as the most frequently used codons, whereas the least-used codons were different in each species and between regional forms of P. westermani. The phylogenetic tree reconstructed from a concatenated alignment of amino acids of 12 PCGs from 36 strains/26 species/5 families of trematodes confirmed that the Paragonimidae is monophyletic, with 100% nodal support. Paragonimus skrjabini miyazakii was resolved as a sister to P. heterotremus. The P. westermani clade was clearly separate from remaining congeners. The latter clade was comprised of 2 subclades, one of the East-Asian and the other of the Indian Type 1 samples. Additional mitogenomes in the Paragonimidae are needed for genomic characterization and are useful for diagnostics, identification and genetic/ phylogenetic/ epidemiological/ evolutionary studies of the Paragonimidae.

Recent and old duplications in crustaceans "Internal Transcribed Spacer 1″: structural and phylogenetic implications.

Internal Transcribed Spacer structures are important in preserving accessibility to specific enzymes for the maturation of rRNAs. ITS1 sequences reported in the literature in Crustaceans range between 182 and 820 bp and are characterized by the absence of repeats or the presence of only a limited number of microsatellites. Here, we sequenced ITS1 for a range of shrimp families (infraorder Caridea) and show that most taxa have much larger ITS1 sequences. We find a high number of microsatellites in Alpheus hebes and Crangon crangon and we report repeat units in Pandalidae, Palaemonidae and mainly in Alpheidae species. Up to four repeats were found in A. vanderbilti (1915 bp), A. rostratus (1635 bp) and A. lottini (1625 bp). In general, four helices were found in ITS1. Repeat units led to extra hairpins and loops. No conserved positions occurred except in helix 4. Three clades were defined in A. lottini for the first time. We estimated the ITS1 divergence rate for the three clades of A. lottini collected in French Polynesia using existing calibrations of substitution rates. Rates of sequence evolution are largely influenced by repeat units, which likely evolve separately. By comparison with COI marker, we estimated the divergence rate of the whole ITS1 sequence to range from 0.5 to 1.4% Pmy and between 0.12 and 0.5% for the 3' end of ITS1 located outside the repeat units. Given the degree of identity between repeats, we suggest that a duplication event recently occurred in A. floridanus (98% identity) whereas an ancient duplication happened in A. sulcatus (50% identity) early at the origination of the group Alpheidae, approximately 50 mya ago. In conclusion, our results highlight an over representation of shorter ITS1 sequences in public repositories, and underlines the importance to further understand patterns of molecular evolution of this functionally important gene.

Analysis of tandem repeat units of the promoter of capsanthin/capsorubin synthase (Ccs) gene in pepper fruit.

Capsanthin/capsorubin synthase (Ccs) gene is a key gene that regulates the synthesis of capsanthin and the development of red coloration in pepper fruits. There are three tandem repeat units in the promoter region of Ccs, but the potential effects of the number of repetitive units on the transcriptional regulation of Ccs has been unclear. In the present study, expression vectors carrying different numbers of repeat units of the Ccs promoter were constructed, and the transient expression of the ß-glucuronidase (GUS) gene was used to detect differences in expression levels associated with the promoter fragments. These repeat fragments and the plant expression vector PBI121 containing the 35s CaMV promoter were ligated to form recombinant vectors that were transfected into Agrobacterium tumefaciens GV3101. A fluorescence spectrophotometer was used to analyze the expression associated with the various repeat units. It was concluded that the constructs containing at least one repeat were associated with GUS expression, though they did not differ from one another. This repeating unit likely plays a role in transcription and regulation of Ccs expression.

Mycobacterial Interspersed Repetitive Unit Can Predict Drug Resistance of Mycobacterium tuberculosis in China.

BACKGROUND: Recently, Mycobacterial Interspersed Repetitive Unit (MIRU) was supposed to be associated with drug resistance in Mycobacterium tuberculosis (M. tuberculosis), but whether the association exists actually in local strains in China was still unknown. This research was conducted to explore that association and the predictability of MIRU to drug resistance of Tuberculosis (TB). METHODS: The clinical isolates were collected and the susceptibility test were conducted with Lowenstein-Jensen (LJ) medium for five anti-TB drug. Based on PCR of MIRU-VNTR (Variable Number of Tandem Repeat) genotyping, we tested the number of the repeat unite of MIRU. Then, we used logistic regression to evaluate the association between 15 MIRU and drug resistance. In addition, we explored the most suitable MIRU locus of identified MIRU loci for drug resistance by multivariate logistic regression. RESULTS: Of the 102 strains, one isolate was resistant to rifampicin and one isolate was resistant to streptomycin. Among these fifteen MIRU, there was a association between MIRU loci polymorphism and anti-tuberculosis drug resistance, ETRB (P = 0.03, OR = 0.19, 95% CI 0.05-0.81) and ETRC (P = 0.01, OR = 0.14, 95% CI 0.03-0.64) were negatively related to isoniazid resistance; MIRU20 (P = 0.05, OR = 2.87, 95% CI 1.01-8.12) was positively associated with ethambutol resistance; and QUB11a (P = 0.02, OR = 0.79, 95% CI 0.65-0.96) was a negative association factor of p-aminosalicylic acid resistance. CONCLUSION: Our research showed that MIRU loci may predict drug resistance of tuberculosis in China. However, the mechanism still needs further exploration.

Rapid detection of structural variation in a human genome using nanochannel-based genome mapping technology.

BACKGROUND: Structural variants (SVs) are less common than single nucleotide polymorphisms and indels in the population, but collectively account for a significant fraction of genetic polymorphism and diseases. Base pair differences arising from SVs are on a much higher order (>100 fold) than point mutations; however, none of the current detection methods are comprehensive, and currently available methodologies are incapable of providing sufficient resolution and unambiguous information across complex regions in the human genome. To address these challenges, we applied a high-throughput, cost-effective genome mapping technology to comprehensively discover genome-wide SVs and characterize complex regions of the YH genome using long single molecules (>150 kb) in a global fashion. RESULTS: Utilizing nanochannel-based genome mapping technology, we obtained 708 insertions/deletions and 17 inversions larger than 1 kb. Excluding the 59 SVs (54 insertions/deletions, 5 inversions) that overlap with N-base gaps in the reference assembly hg19, 666 non-gap SVs remained, and 396 of them (60%) were verified by paired-end data from whole-genome sequencing-based re-sequencing or de novo assembly sequence from fosmid data. Of the remaining 270 SVs, 260 are insertions and 213 overlap known SVs in the Database of Genomic Variants. Overall, 609 out of 666 (90%) variants were supported by experimental orthogonal methods or historical evidence in public databases. At the same time, genome mapping also provides valuable information for complex regions with haplotypes in a straightforward fashion. In addition, with long single-molecule labeling patterns, exogenous viral sequences were mapped on a whole-genome scale, and sample heterogeneity was analyzed at a new level. CONCLUSION: Our study highlights genome mapping technology as a comprehensive and cost-effective method for detecting structural variation and studying complex regions in the human genome, as well as deciphering viral integration into the host genome.

C9ORF72 hexanucleotide repeat number in frontotemporal lobar degeneration: a genotype-phenotype correlation study.

Expansion of a hexanucleotide repeat in the C9ORF72 gene has been identified as the most common pathogenic mutation in families with autosomal dominant frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis. Herein we investigated frequency and penetrance of the C9ORF72 hexanucleotide repeat pathological expansion in a large cohort of familial and sporadic FTLD and related disorders (FTLD and related disorders, n = 388; Controls, n = 201). Moreover, we weighed the impact of C9ORF72 genotype on clinical phenotype taking into account the hexanucleotide repeat units number as a possible disease modifier. In our cohort, the C9ORF72 pathological expansion: (i) showed a prevalence of 7.5%; (ii) showed a full penetrance by the age of 80; (iii) was rarely found in sporadic patients; (iv) was solely associated with FTLD; (v) was mainly associated with bvFTD clinical subtype; and (vi) was associated with earlier age of onset in the youngest generation compared with the previous generation within a pedigree. Interestingly, intermediate C9ORF72 expansion had a risk effect in familial/sporadic FTLD. Eventually, the C9ORF72 repeat units number influenced the disease phenotype in terms of age of onset and associated clinical subtype. Genome-wide studies in well characterized clinical cohorts will be essential in order to decipher pathways of disease expression in C9ORF72-associated neurodegeneration.

Study on the molecular size distribution and the structural characteristics of group B meningococcal cap-sular polysaccharides / 中华微生物学和免疫学杂志

Objective To investigate the molecular size distribution and the structure of group B me-ningococcal capsular polysaccharides for the development of vaccines .Methods The molecular size distribution of group B meningococcal capsular polysaccharides was analyzed by chromatography on a Sepharose CL -4B col-umn.The molecular weight of repeat units were measured by matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS).The structural characteristics of group B meningococcal capsular polysaccharides were analyzed by nuclear magnetic resonance ( NMR) based on the chemical shift of all charac-teristic protons by using group C meningococcal capsular polysaccharides and sialic acid as the controls .Results The KD value of group B meningococcal capsular polysaccharides extracted from 15 strains were ranged from 0.60 to 0.76.The molecular weight of repeat units was 284, which was identical to the theoretical value .The group B meningococcal capsular polysaccharides were 2→8 linked homopolymers of sialic acid lacking O-acetyl groups.Conclusion The group B meningococcal capsular polysaccharides had lower molecular weights , which might result in their poor immunogenicity .The structure of group B meningococcal capsular polysaccharides could be quickly and accurately analyzed by NMR technology .

Enterobacterial small mobile sequences carry open reading frames and are found intragenically--evolutionary implications for formation of new peptides.

Intergenic repeat units of 127-bp (RU-1) and 168-bp (RU-2), as well as a newly-found class of 103-bp (RU-3), represent small mobile sequences in enterobacterial genomes present in multiple intergenic regions. These repeat sequences display similarities to eukaryotic miniature inverted-repeat transposable elements (MITE). The RU mobile elements have not been reported to encode amino acid sequences. An in silico approach was used to scan genomes for location of repeat units. RU sequences are found to have open reading frames, which are present in annotated gene loci whereby the RU amino acid sequence is maintained. Gene loci that display repeat units include those that encode large proteins which are part of super families that carry conserved domains and those that carry predicted motifs such as signal peptide sequences and transmembrane domains. A putative exported protein in Y. pestis and a phylogenetically conserved putative inner membrane protein in Salmonella species represent some of the more interesting constructs. We hypothesize that a major outcome of RU open reading frame fusions is the evolutionary emergence of new proteins.