Repair of Rat Calvarial Bone Defect by Using Exosomes of Umbilical Cord-Derived Mesenchymal Stromal Cells Embedded in Chitosan/Hydroxyapatite Scaffolds.

PURPOSE: To seek out the bone regeneration effect of human umbilical cord-mesenchymal stem cell (hUC-MSC)-derived exosomes of loaded chitosan/hydroxyapatite (CS/HA) scaffold in a rat calvarium bone regeneration model. MATERIALS AND METHODS: The hUC-MSC exosomes were purified and characterized. The scaffolds were prepared by a freeze-drying method. Animals were divided into five groups, and the CS/HA/exosome (CS/HA/Exo) scaffolds were transplanted to 5 × 2-mm critical-sized calvarial bone defects for repair in rats. All animals were sacrificed at the postoperative sixth week. Immunohistochemical and histologic analyses were performed. RESULTS: Scanning electron microscopy (SEM) images showed that the exosomes were round-shaped vesicles with bounded membrane, and the diameter of the exosomes was 83.728 ± 27.269 nm. Histologic analysis showed that mean new bone volumes were statistically significantly higher in the CS/HA/Exo group (1.83 ± 0.54, PCS/Exo-CS/HA/Exo = .000), and other new bone volumes in the other groups were statistically significant compared with the control (CS/Exo 1.50 ± 0.14 mm3; CS 1.20 ± 0.43 mm3; control 1.06 ± 0.10 mm3; and CS/HA 1.43 ± 0.66 mm3). CONCLUSION: The CS/HA/Exo combination is a novel treatment for bone defect repair to induce bone formation. The CS scaffold can significantly promote bone regeneration compared with the control. Moreover, the combination with HA and exosomes is promising for applications in bone tissue regeneration.

The first detection of betanodavirus reassortant genotype (RGNNV/SJNNV) isolated from gilthead sea bream (Sparus aurata) in the Turkish coastlines: The importance of screening and monitoring studies for identifying the source of the infection.

Viral nervous necrosis (VNN) is now endemic in the Mediterranean basin and the RGNNV genotype betanodavirus has caused frequent epidemics in European sea bass for a long time. Unexpected and increasing VNN epidemics have been reported in gilthead sea bream (GSB) farms in the last few years, from which the RGNNV/SJNNV genotype has been mostly isolated. The aim of this study was to perform a molecular characterization of the betanodavirus isolated from GSB (weighing 90-100 g) in a marine fish farm in the Aegean Sea and also, as an early warning exercise, to investigate the presence/absence of the virus in associated nearby farms (n:20) and in hatcheries (n:3). No virus was detected in any of the nearby farms or two hatcheries. However, in one hatchery, betanodavirus was detected in a 160-day-old GSB. The identified betanodavirus was genotyped as reassortant RGNNV/SJNNV and was phylogenetically related to the virus detected in the farm located in the Aegean sea. There have been multiple detections of the RGNNV genotype in Turkish coastal waters; however, the RGNNV/SJNNV genotype has been detected for the first time and it should be an early warning to focus attention on betanodaviruses in Turkish aquaculture.

Determination of VP2 sequence-based virulence motifs and phylogenetic analysis of domestic Turkish IPNV isolates.

Infectious pancreatic necrosis (IPN) is a highly contagious disease of young salmonid fish and is one of the most severe economic diseases in aquaculture. In Turkey, an increase in infectious pancreatic necrosis virus (IPNV) outbreaks in freshwater rainbow trout have been reported in recent years. This study aimed to analyze the VP2 gene from recent IPNV isolates from Turkey to determine whether there are epidemiological links between IPNV isolates from rainbow trout (Oncorhynchus mykiss; 62) and sea bass (Dicentrarchus labrax; 1), wild turbot (Scophthalmus maximus; 1) and the environment in order to investigate potential wild and farmed fish interactions. In this study, 62 Turkish IPNV isolates collected over 10 years (2005-2014) from rainbow trout, sea bass and turbot were genotypically characterized. The phylogenetic analysis indicated that Turkish IPNV isolates are closely related to strains from Denmark, Iran and Spain and that all Turkish IPNV isolates belong to genogroup V, serotype A2 (Sp strain). Furthermore, low genetic diversity was found among the Turkish isolates (identity, 95.5%-100% nucleotides and 97.8%-100% amino acids). The result of the analysis of the amino acid residues found at positions 217, 221 and 247 (proline, threonine and alanine, respectively) could be associated with virulence.

Inactivated infectious pancreatic necrosis virus (IPNV) vaccine and E.coli-expressed recombinant IPNV-VP2 subunit vaccine afford protection against IPNV challenge in rainbow trout.

Infectious pancreatic necrosis (IPN) is a highly contagious disease causing high mortality in juvenile trouts. Since there is no effective way to treatment against IPNV, early diagnosis and prevention play an important role in combating the disease. The different types of IPNV vaccines (inactive, live, recombinant, DNA, etc) have been produced from local isolates and have been used in developed countries. In Turkey, there is no commercial licensed vaccines against IPNV. Due to this reason, IPNV vaccine is needed in Turkey. The production of recombinant VP2 subunit vaccine (IPNV-VP2) and inactivated whole particle virus vaccine (IPNV-WPV) were attempted from selected isolate belong to sp serotype. For this purpose; the virus was produced in RTG-2 cell line and RT-PCR amplification was performed by using primers with restriction enzymes. The whole VP2 gene was cloned into a plasmid vector and VP2 was expressed by using E. coli expression system. A trial was conducted to determine the immunity ability of IPNV-VP2 and IPNV-WPV in rainbow trout. According to the SN50 assay, the IPNV-WPV stimulates immune response faster than the IPNV-VP2 vaccine. Besides, the relative percent of Survive (RPS) was detected as 79% in fish vaccinated with IPNV-WPV and 70% in fish vaccinated with IPNV-VP2. Thus, we can say that the recombinant vaccine of IPNV-VP2 is almost protected against IPNV infection as well as the inactive vaccine.

Detection of RGNNV genotype betanodavirus in the Black Sea and monitoring studies.

Viral nervous necrosis (VNN), caused by betanodavirus, is a significant viral infection that threatens marine aquaculture. Freshwater and marine fish farms in Turkey are subjected to annual pathogen screenings. In 2016, during the Nervous Necrosis Virus screening program conducted in the Black Sea, betanodavirus was unexpectedly detected using real-time reverse transcription-polymerase chain reaction in apparently healthy sea bass. Phylogenetic analysis of both the RNA1 and RNA2 segments of the virus determined that the betanodavirus detected was red-spotted grouper nervous necrosis virus genotype (RGNNV). Following the initial discovery of betanodavirus in the Black Sea, monitoring studies performed over a 3 yr period have not indicated any additional presence of the virus. The absence of clinical symptoms related to VNN disease in the area's marine fish farms and the surrounding detection zone, and the fact that the virus has not been detected anew in monitoring programmes conducted following the initial detection, indicate that there is no virus circulation in the detection zone.