Synthesis and characterizations of natural limestone-derived nano-hydroxyapatite (HAp): a comparison study of different metals doped HAps on antibacterial activity

Earth-abundant mineral limestone obtained from North Sumatera, Indonesia, has been utilized to synthesize nano-hydroxyapatite (HAp). Although HAp is biocompatible to the human bone, its antibacterial activity is still very low. Herein, different metal ions (i.e., Ag, Cu, Zn, and Mg) were doped into HAp to improve the antibacterial activity. The as-synthesized HAp was characterized by X-ray ray diffraction (XRD), field-emission scanning electron microscope (FE-SEM), energy disperse spectroscopy (EDS), Fourier transmission infrared spectroscopy (FTIR), X-ray photoelectron spectroscopy (XPS), and Brunauer–Emmett–Teller (BET). The antibacterial test showed that the performance of HAp to inactivate bacterial growth was significantly improved after incorporating the metal ion dopants into HAp. Ag-HAp exhibited the highest activity toward E. coli and S. aureus with an antibacterial rate of 99.9 ± 0.1%, followed by Zn-HAp, Cu-HAp, and Mg-HAp.

Antibacterial activities of different metal ion doped HAp towards (a) E. coli and (b) S. aureus bacteria.     

Diarrhoea episodes and treatment-seeking behaviour in a slum area of North Jakarta, Indonesia

Visits to household during a census in an impoverished area of north Jakarta were used for exploring the four-week prevalence of diarrhoea, factors associated with episodes of diarrhoea, and the patterns of healthcare use. For 160,261 urban slum-dwellers, information was collected on the socioeconomic status of the household and on diarrhoea episodes of individual household residents in the preceding four weeks. In households with a reported case of diarrhoea, the household head was asked which form of healthcare was used first. In total, 8,074 individuals (5%)--13% of children aged less than five years and 4% of adults--had a diarrhoea episode in the preceding four weeks. The two strongest factors associated with a history of diarrhoea were a diarrhoea episode in another household member in the four weeks preceding the interview (adjusted odds ratio [OR] 11.1; 95% confidence interval [CI] 10.4-11.8) and age less than five years (adjusted OR 3.4; 95% CI 3.2-3.5). Of the 8,074 diarrhoea cases, 1,969 (25%) treated themselves, 1,822 (23%) visited a public-health centre (PHC), 1,462 (18%) visited a private practitioner or a private clinic, 1,318 (16%) presented at a hospital, 753 (9%) bought drugs from a drug vendor, and 750 (9%) used other healthcare providers, such as belian (traditional healers). Children with diarrhoea were most often brought to a PHC, a private clinic, or a hospital for treatment. Compared to children, adults with diarrhoea were more likely to treat themselves. Individuals from households in the lowest-income group were significantly more likely to attend a PHC for treatment of diarrhoea compared to individuals from households in the middle- and higher-income groups.     

Development of an entirely plasmid-based reverse genetics system for 12-segmented double-stranded RNA viruses

The family Reoviridae is a nonenveloped virus group with a double-stranded (ds) RNA genome comprising 9 to 12 segments. In the family Reoviridae, the genera Cardoreovirus, Phytoreovirus, Seadornavirus, Mycoreovirus, and Coltivirus contain virus species having 12-segmented dsRNA genomes. Reverse genetics systems used to generate recombinant infectious viruses are powerful tools for investigating viral gene function and for developing vaccines and therapeutic interventions. Generally, this methodology has been utilized for Reoviridae viruses such as Orthoreovirus, Orbivirus, Cypovirus, and Rotavirus, which have genomes with 10 or 11 segments, respectively. However, no reverse genetics system has been developed for Reoviridae viruses with a genome harboring 12 segments. Herein, we describe development of an entire plasmid-based reverse genetics system for Tarumizu tick virus (TarTV) (genus Coltivirus, family Reoviridae), which has a genome of 12 segments. Recombinant TarTVs were generated by transfection of 12 cloned complementary DNAs encoding the TarTV genome into baby hamster kidney cells expressing T7 RNA polymerase. Using this technology, we generated VP12 mutant viruses and demonstrated that VP12 is an N-glycosylated protein. We also generated a reporter virus expressing the HiBiT-tagged VP8 protein. This reverse genetics system will increase our understanding of not only the biology of the genus Coltivirus but also the replication machinery of the family Reoviridae.

The family Reoviridae is a nonenveloped virus group classified into 15 genera. These viruses have double-stranded (ds) RNA genomes with 9 to 12 segments. This family includes several important pathogens in both humans and animals. Mammalian orthoreovirus (MRV) and Nelson Bay orthoreovirus (NBV), which belong to the genus Orthoreovirus, have genomes consisting of 10 segments of dsRNA. MRV is an experimental model for studies of Reoviridae virus replication and pathogenesis. NBV, classified into the fusogenic subgroup of this genus, is associated with acute respiratory tract infections in humans (1–3). The fusogenic orthoreoviruses encode a unique fusion-associated small transmembrane (FAST) protein associated with cell–cell fusion and viral pathogenesis (4–6). Bluetongue virus (BTV) and African horse sickness virus (AHSV) belong to the genus Orbivirus, have genomes with 10 segments of dsRNA, and cause severe diseases in domestic animals (7, 8). Rotavirus (RV, genus Rotavirus) has a genome with 11 segments of dsRNA and causes severe diarrhea in young children. RV infection is responsible for 128,500 deaths per year worldwide, predominately in developing countries (9). Colorado tick fever virus (CTFV), an arthropod-borne virus transmitted by ticks and belonging to the genus Coltivirus, has a dsRNA genome comprising 12 segments. CTFV causes a variety of symptoms in humans, including abrupt fever, chills, headache, myalgia, and abdominal pain (10, 11). Within the genus Coltivirus (in addition to CTFV), Eyach virus, Shelly Headland virus, Kundal virus, and Tarumizu tick virus (TarTV) have been isolated from, or detected in, ticks in Europe, Australia, India, and Japan, respectively (12–17). Taï Forest reovirus was isolated from free-tailed bats in Côte d’Ivoire (18), and Lishui pangolin virus was detected in pangolins in China (19). These reports suggest that coltiviruses are distributed in multiple species worldwide. However, the molecular mechanism underlying the propagation and pathogenesis of these viruses remains largely unknown.Reverse genetics systems are powerful tools used to study many aspects of viral biology and virus–host interactions and also provide an opportunity to generate recombinant viruses that can be used for vaccines or as viral vectors. This technology has been used for several viruses in the family Reoviridae. In the genus Orthoreovirus, an entirely plasmid-based reverse genetics system was developed for MRV in 2007 (20). This is the first example of engineering recombinant Reoviridae viruses entirely from cloned complementary DNAs (cDNAs). This system was established by cotransfection of cloned cDNAs representing 10 MRV gene segments, each flanked by the T7 promoter and hepatitis delta virus (HDV) ribozyme, into cells expressing T7 RNA polymerase. Subsequently, a reverse genetics system for NBV, belonging to a fusogenic reovirus group, was established based on the MRV rescue system (21). In the genus Orbivirus, RNA- and DNA-based reverse genetics systems were developed for BTV, AHSV, and Epizootic hemorrhagic disease virus (22–25). Although development of a reverse genetics system for the genus Rotavirus has lagged behind those for the genera Orthoreovirus and Orbivirus, our group recently developed the first plasmid-based reverse genetics system for RV by cotransfecting plasmids encoding unique heterogeneous viral proteins, NBV FAST, and vaccinia virus capping enzyme as rescue enhancers along with the 11 RV T7 promoter-based rescue plasmids (26). In addition, reverse genetics systems were also developed for Bombyx mori cypovirus and Dendrolimus punctatus cypovirus (genus Cypovirus), which are insect pathogens and have a genome consisting of 10 segments of dsRNA (27, 28). Since reverse genetics systems were developed for Reoviridae viruses with genomes containing 10 or 11 segments of dsRNA, studies of the family Reoviridae have advanced markedly. However, to date, no reverse genetics system has been established for Reoviridae viruses with 12 genome segments.In this study, we isolated a TarTV strain from a raccoon dog postmortem. Using this TarTV strain, we established an entire plasmid-based reverse genetics system and rescued VP12 mutant viruses and a HiBiT-tagged reporter virus. This system is a useful tool to generate recombinant coltiviruses with 12 dsRNA genome segments.     

Correction: The burden of diarrhoea, shigellosis, and cholera in North Jakarta, Indonesia: findings from 24 months surveillance

This is a correction of an earlier published article.     

Osteotome sinus floor elevation technique without grafting material: 3-year results of a prospective pilot study

Objective: In a prospective pilot study, short≤10 mm ITI-SLA implants were placed in the resorbed posterior maxilla by means of an osteotome sinus floor elevation (OSFE) procedure without grafting material. This paper presents 3-year data assessing bone-level changes around implants.
Material and methods: Twenty-five implants were placed in 17 patients to rehabilitate 16 molar and nine premolar sites. The mean residual bone height (RBH) was 5.4 ± 2.3 mm. A healing period of 3–4 months was allowed before abutment tightening. Most implants (21/25) were 10 mm long, and the others were 8 and 6 mm long. At the 3-year control, endo-sinus bone gain, implant length protruding into the sinus and crestal bone loss (CBL) were measured on periapical radiographs.
Results: All implants fulfilled the survival criteria. Despite the absence of grafting material, implants were embedded into newly formed bone tissue. All implants gained endo-sinus bone; the mean gained bone was 3.1 ± 1.5 mm. The residual protrusion length decreased from 4.9 ± 2.1 to 1.8 ± 2.1 mm. CBL was 0.9 ± 0.8 mm.
Conclusions: This study confirms that the OSFE procedure without grafting material is sufficient to create bone beyond the natural limit of the sinus. On the mid-term of 3 years, the technique was found to be predictable in the posterior maxilla when the RBH is limited. Implants gained endo-sinus bone despite the lack of grafting material. Bone gain was still improving over the first-year control. No shrinkage of the augmented area was observed.     

Dillenia suffruticosa exhibited antioxidant and cytotoxic activity through induction of apoptosis and G2/M cell cycle arrest

Ethnopharmacological relevance

Dillenia suffruticosa (Family: Dilleniaceae) locally known as Simpoh air has been reported to be used traditionally to treat cancerous growth. Therefore, the present study was attempted to investigate the antioxidant and cytotoxic properties of different parts (root, flower, fruit and leaf) of D. suffruticosa extracts.

Methods and materials

In this study, direct solvent extraction (aqueous and methanol) from different parts of D. suffruticosa (root, flower, fruit and leaf) were carried out. Antioxidant activities of D. suffruticosa extract were determined by using DPPH, ABTS FRAP and β-carotene bleaching assays. Cytotoxicity and cell cycle arrest of the active extract were determined using MTT assay and flow cytometer, respectively. Sequential solvent extraction (hexane, DCM, EtOAc, and MeOH) were also carried out in root of D. suffruticosa to further evaluate the antioxidant and cytotoxic activity of the different solvent extracts.

Results

Methanol (MeOH) root extract showed the highest TPC, antioxidant and cytotoxic activities (especially towards HeLa) compared to others (P<0.05). Based on the results, sequential solvent extraction (hexane, DCM, EtOAc and MeOH) was carried out in the roots of D. suffruticosa. MeOH extract exhibited the highest antioxidant activities among others and significantly correlated (P<0.05) with TPC, suggesting the important contribution of phenolic compounds to its antioxidant activity. On the other hand, the DCM and EtOAc exhibited higher cytotoxic activity to selected cancer cells (HeLa, MCF-7, MDA-MB-231, A549 and HT29) compared to others. In short, there is no established correlation between antioxidant and cytotoxic activities of D. suffruticosa extracts indicating that an agent with high antioxidant activities will not necessarily possesses good cytotoxic activities in return. Qualitative phytochemical screening of D. suffruticosa extracts suggested the presence of saponins, triterpenes, sterols, and polyphenolic compounds which are believed to contribute to the cytotoxic activities.

Conclusion

It is suggested that the cytotoxicity of the active extracts in HeLa was due to the induction of apoptosis and cell cycle arrest at G₂/M.