Characterization of the complete mitochondrial genomes of two Ixodes ticks, I. nipponensis and Ixodes (Pholeoixodes) sp.

In this study, the authors sequenced and characterized the complete mitochondrial (mt) genomes of two hard ticks of the genus Ixodes, I. nipponensis and Ixodes (Pholeoixodes) sp., which were 14 505 and 14 543 bp in length, respectively. Their mt genomes encoded 37 genes, including 13 protein-coding genes (PCGs), 22 transfer RNA genes and two ribosomal RNA genes, and have only one non-coding region. The gene order in their mt genomes was the same as that of other Ixodes spp. mt genomes. The average sequence identity, combined nucleotide diversity, non-synonymous/synonymous substitutions ratio analyses consistently demonstrated that cox1, rrnS, cox2, cox3 and cytb were the most conserved and atp8, nad6 and nad2 were the most variable genes across Ixodes mitogenomes. Phylogeny of the present Ixodes spp., and other selected hard tick species, based on concatenated amino acid sequences of PCGs, confirmed their position within the genus Ixodes and sub-family Ixodinae. The novel mt markers described herein will be useful for further studies of the population genetics, molecular epidemiology and systematics of hard ticks.

Comparative analysis of microbial community in the whole body and midgut from fully engorged and unfed female adult Melophagus ovinus.

Melophagus ovinus is a type of ectoparasite infesting sheep. Data regarding the comprehensive bacterial community associated with the whole body and midgut of M. ovinus under different engorged statuses are required. Melophagus ovinus were collected from the city of Jiuquan, China. Bacterial DNA was extracted from the whole body and midgut of fully engorged female adults, or newly hatched and unfed adult female M. ovinus. The 16S rRNA gene V3-V4 hypervariable regions were sequenced using the IonS5™XL platform (Thermo Fisher Scientific, Waltham, MA, U.S.A.). The whole body bacterial diversity of the newly hatched, unfed adult females was greater compared with that of the other three samples. Proteobacteria was the dominant bacterial phylum in all of the samples. Of the 42 total bacterial genera present in all of the experimental samples, Arsenophonus, Bartonella and Wolbachia were the dominant genera. The relative abundance of Arsenophonus in midgut was greater than that in the whole body. The relative abundance of Bartonella in fully engorged adults was far greater than those in newly hatched, unfed adults. The relative abundance of Wolbachia was highest in the whole body of newly hatched, unfed adults. Seventeen bacterial species were identified in all experimental samples. Bartonella chomelii, Streptococcus hyointestinalis and Escherichia coli were the first species reported in M. ovinus.

Cloning of four HSPA multigene family members in Haemaphysalis flava ticks.

The heat shock protein 70 (HSPA) family and their genes have been studied in ticks and are considered as possible antigen candidates for the development of anti-tick vaccines. However, knowledge about their members, structure and function in ticks is incomplete. Based on our transcriptomic data, the full length of four HSPA genes in Haemaphysalis flava (Acari: Ixodidae) was cloned via rapid amplification of cDNA ends. The open reading frame of HSPA2A, HSPA2B, HSPA5 and HSPA9 was 1920, 1911, 1983 and 2088 bp in length, respectively. Three family signatures and one localization motif were in the encoding proteins. HSPA2A and HSPA2B were predicted to be located at cytoplasm/nucleus, whereas HSPA5 and HSPA9 were at endoplasmic reticulum and mitochondria, respectively. In silico simulation demonstrated that those proteins had distinct numbers of α-helixes, extended strands and coils, and different antigenic epitopes. Expression of HSPA5 and HSPA9 in the salivary gland was significantly higher in partially-engorged female adult ticks than the fully-engorged (P < 0.01) as shown by a quantitative polymerase chain reaction. Our data indicated that H. flava ticks had at least four HSPA genes encoding proteins with different cellular locations, structures and expression profiles, suggesting their diverse roles in tick biology.

Mitochondrial genome of Amblyomma javanense: a hard tick parasite of the endangered Malayan pangolin (Manis javanica).

Amblyomma javanense is an important ectoparasite of Manis javanica, although the population genetics, molecular biology and systematics of A. javanense remain poorly understood. In the present study, the mitochondrial genome of A. javanense was sequenced using the Illumina HiSeq sequencing platform (Illumina, San Diego, CA, U.S.A.) and compared with the genomes of two closely related species: Amblyomma fimbriatum and Amblyomma americanum. The intraspecies and interspecies relationships of A. javanense and another 21 selected species were investigated by constructing a maximum-likelihood tree and a neighbour-joining tree. The mitochondrial genome of A. javanense was 14 780 bp in length and contained 13 protein-coding genes, 22 transfer RNA genes, two ribosomal RNA genes and two control regions. The results of the comparisons indicate that there is great similarity among these three species, and both trees indicate that A. javanense is a member of the Amblyomminae. The study of A. javanense of pangolins also indicates the premise and foundation of the relationship between the parasite and other species.

[Neocartilage formation in vitro using transduced mesenchymal stem cells cultured on biomimetic biodegradable polymer scaffolds].

OBJECTIVE: To investigate the effect of transforming growth factor (TGF) beta 1 gene transfection on the growth of mesenchymal stem cells(MSCs) and to evaluate a new biomimetic biodegradable polymer as scaffolds for applications in articular cartilage tissue engineering. METHODS: Principles of tissue engineering were combined organically with principles of gene therapy to produce cultured periosteum-derived MSCs transduced with the full-length rat TGF-beta 1 cDNA in vitro. These cells were then seeded onto three-dimensional porous poly-DL-lactide scaffolds modified with poly-L-lysine that mimicked cell-binding domains found on natural extracellular matrix to promote specific cell adhesion. The adhesion, proliferation, and differentiation of the transfected MSCs were examined with scanning electron microscope within 2 weeks. RESULTS: All cells adhered to the biomimetic matrices well, but more cartilage-like tissue was formed for TGF-beta 1 gene modified MSCs/scaffolds composites than for the control groups. Transfer of gene encoding TGF-beta 1 to MSCs promoted its proliferation and differentiation significantly. CONCLUSIONS: The TGF-beta 1 gene transduced MSCs/biomimetic matrix composites used in this study was the first attempt to apply the principles of molecular tissue engineering for articular cartilage repair. This new molecular tissue engineering approach could be of potential benefit to repair damaged articular cartilage, especially in osteoarthritis. The new biomimetic biodegradable polymer matrices modified with biomolecules not only have good structural compatibility, but also have better interfacial compatibility and bioactivity, and can be used as scaffolds for articular cartilage tissue engineering.

Sustained outward current observed after I(to1) inactivation in rabbit atrial myocytes is a novel Cl- current.

In rabbit atrial myocytes, depolarization of the membrane results in a rapidly activating transient outward current (I(to)) that then decays to a sustained level. The sustained current (Isus) remains constant for at least 5 s during continued depolarization. The present study was designed to identify the ionic mechanism underlying Isus with the use of whole cell voltage-clamp techniques. After exposure to 2 mM 4-aminopyridine (4-AP), the 4-AP-sensitive transient outward current (I(to1)) was abolished, but Isus was unaffected. Isus was not blocked by the K+ channel blockers tetraethylammonium chloride and Ba2+, was not changed by increasing superfusate K+ concentration, and was still present when K+ was replaced by Cs+ in both the superfusate and the pipette. Isus was significantly reduced by the Cl- transport blockers 4-acetamido-4'-isothiocyanatostilbene-2.2'-disulfonic acid and 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid. The current-voltage relations of Isus showed outward rectification, and the reversal potential of Isus shifted with changes in the transmembrane Cl- gradient in the fashion expected for a Cl- current. We conclude that Isus in rabbit atrium is due to a noninactivating Cl- current which, unlike previously described cardiac Cl- currents, is manifest in the absence of exogenous stimulators of adenosine 3',5'-cyclic monophosphate formation, cytosolic Ca2+ transients, or cell swelling.