Barriers to Childhood Immunisation and Local Strategies in Four Districts in South Africa: A Qualitative Study.

Introduction: In South Africa over the past 20 years, immunisation has saved countless lives as well as prevented illnesses and disabilities. Despite this, vaccine-preventable illnesses remain a danger. The demand for and uptake of immunisation services are shaped by a variety of factors that can either act as barriers or facilitators to immunisation uptake. The aim of this project was to identify the supply and demand barriers and develop local strategies to improve childhood immunisation in four zero-dose districts in South Africa. Materials and Methods: This study used a mixed-method approach. In each of these four districts, 15 in-depth key informant interviews with health workers and local health managers and four focus group discussions (10 participants per focus group discussion) with community members and caregivers were held over a three-month period. Transcribed interviews were thematically analysed using qualitative analysis software (Nvivo®) into 10 factors as identified as important in influencing immunisation demand and uptake in previous studies. A further four were identified during the data analysis process. Results: Despite the varying role of factors affecting demand and uptake of immunisation services, three consistent findings stand out as major barriers across all districts. The first is interaction with healthcare staff. This clearly highlights the crucial role that the interactions between patients and staff play in shaping perceptions and behaviours related to immunisation services. The second is the overall experience of care at healthcare facilities. This emphasises the role that patient experience of services plays in perceptions and behaviours related to immunisation services. The third is family dynamics. This highlights the important role family dynamics play in shaping individuals' decisions regarding immunisation uptake as well as the impact it has on the ability of people to access health services. Discussion: The role played by the different factors in the demand and uptake of immunisation services varied across the four districts examined in this study. Each of the districts presents a unique landscape where different factors have varying degrees of importance in affecting the utilisation of immunisation services. In some districts, certain factors are major barriers, clearly hindering the demand and uptake of immunisation services, while in others, these same factors might be a relatively minor barrier. This discrepancy highlights the unique nature of healthcare challenges across the districts and the need for tailored strategy recommendations to address them effectively.

Transient Retrovirus-Based CRISPR/Cas9 All-in-One Particles for Efficient, Targeted Gene Knockout.

The recently discovered CRISPR/Cas9 system is widely used in basic research and is a useful tool for disease modeling and gene editing therapies. However, long-term expression of DNA-modifying enzymes can be associated with cytotoxicity and is particularly unwanted in clinical gene editing strategies. Because current transient expression methods may still suffer from cytotoxicity and/or low efficiency, we developed non-integrating retrovirus-based CRISPR/Cas9 all-in-one particles for targeted gene knockout. By redirecting the gammaretroviral packaging machinery, we transiently delivered Streptococcus pyogenes Cas9 (SpCas9) mRNA and single-guide RNA transcripts into various (including primary) cell types. Spatiotemporal co-delivery of CRISPR/Cas9 components resulted in efficient disruption of a surrogate reporter gene, as well as functional knockout of endogenous human genes CXCR4 and TP53. Although acting in a hit-and-run fashion, knockout efficiencies of our transient particles corresponded to 52%-80% of those obtained from constitutively active integrating vectors. Stable SpCas9 overexpression at high doses in murine NIH3T3 cells caused a substantial G0/G1 arrest accompanied by reduced cell growth and metabolic activity, which was prevented by transient SpCas9 transfer. In summary, the non-integrating retrovirus-based vector particles introduced here allow efficient and dose-controlled delivery of CRISPR/Cas9 components into target cells.

A continuous single organic phase process for the lipase catalyzed synthesis of peroxy acids increases productivity.

The design of an optimal process is particularly crucial when the reactants deactivate the biocatalyst. The reaction cascades of the chemo-enzymatic epoxidation where the intermediate peroxy acid is produced by an enzyme are still limited by enzyme inhibition and deactivation by hydrogen peroxide. To avoid additional effects caused by interfaces (aq/org) and to reduce the process limiting deactivation by the substrate hydrogen peroxide, a single-phase concept was applied in a fed-batch and a continuous process (stirred tank), without the commonly applied addition of a carrier solvent. The synthesis of peroxyoctanoic acid catalyzed by Candida antarctica lipase B was chosen as the model reaction. Here, the feasibility of this biocatalytic reaction in a single-phase system was shown for the first time. The work shows the economic superiority of the continuous process compared to the fed-batch process. Employing the fed-batch process reaction rates up to 36 mmol h-1 per gramcat, and a maximum yield of 96 % was achieved, but activity dropped quickly. In contrast, continuous operation can maintain long-term enzyme activity. For the first time, the continuous enzymatic reaction could be performed for 55 h without any loss of activity and with a space-time yield of 154 mmol L-1 h-1, which is three times higher than in the fed-batch process. The higher catalytic productivity compared to the fed-batch process (34 vs. 18 gProd g-1 cat) shows the increased enzyme stability in the continuous process.

Use of bioreactors in maxillofacial tissue engineering.

Engineering of various oral tissues is a challenging issue in contemporary maxillofacial reconstructive research. In contrast to the classic biomaterial approach, tissue engineering is based on the understanding of cell driven tissue formation, and aims to generate new functional tissues, rather than just to implant non-living space holders. Researchers hope to reach this goal by combining knowledge from biology, physics, materials science, engineering, and medicine in an integrated manner. Several major technical advances have been made in this field during the last decade, and clinical application is at the stage of first clinical trials. A recent limitation of extracorporally engineered cellular substitutes is the problem of growing enlarged tissues ex vivo. One of the main research topics is therefore to scale up artificial tissue constructs for use in extended defect situations. To overcome the monolayer inherent two-dimensional cell assembly, efforts have been made to grow cells in a three-dimensional space. Bioreactors have therefore been in focus for a considerable time to build up enlarged tissues. The shift from the ex vivo approach of cell multiplication to the generation of a real tissue growth is mirrored by the development of bioreactors, enabling scientists to grow more complex tissue constructs. This present review intends to provide an overview of the current state of art in maxillofacial tissue engineering by the use of bioreactors, its limitations and hopes, as well as the future research trends.

Principles of cartilage tissue engineering in TMJ reconstruction.

Diseases and defects of the temporomandibular joint (TMJ), compromising the cartilaginous layer of the condyle, impose a significant treatment challenge. Different regeneration approaches, especially surgical interventions at the TMJ's cartilage surface, are established treatment methods in maxillofacial surgery but fail to induce a regeneration ad integrum. Cartilage tissue engineering, in contrast, is a newly introduced treatment option in cartilage reconstruction strategies aimed to heal cartilaginous defects. Because cartilage has a limited capacity for intrinsic repair, and even minor lesions or injuries may lead to progressive damage, biological oriented approaches have gained special interest in cartilage therapy. Cell based cartilage regeneration is suggested to improve cartilage repair or reconstruction therapies. Autologous cell implantation, for example, is the first step as a clinically used cell based regeneration option. More advanced or complex therapeutical options (extracorporeal cartilage engineering, genetic engineering, both under evaluation in pre-clinical investigations) have not reached the level of clinical trials but may be approached in the near future. In order to understand cartilage tissue engineering as a new treatment option, an overview of the biological, engineering, and clinical challenges as well as the inherent constraints of the different treatment modalities are given in this paper.