The present study investigated the influence of pore size of strut-based Diamond and surface-based Gyroid structures for their suitability as medical implants. Samples were made additively from laser powder bed fusion process with a relative density of 0.3 and pore sizes ranging from 300 to 1300 μm. They were subsequently examined for their manufacturability and mechanical properties. In addition, non-Newtonian computational fluid dynamics and discrete phase models were conducted to assess pressure drop and cell seeding efficiency. The results showed that both Diamond and Gyroid had higher as-built densities with smaller pore sizes. However, Gyroid demonstrated better manufacturability as its relative density was closer to the as-designed one. In addition, based on mechanical testing, the elastic modulus was largely unaffected by pore size, but post-yielding behaviors differed, especially in Diamond. High mechanical sensitivity in Diamond could be explained partly by Finite Element simulations, which revealed stress localization in Diamond and more uniform stress distribution in Gyroid. Furthermore, we defined the product of the normalized specific surface, normalized pressure drop, and cell seeding efficiency as the indicator of an optimal pore size, in which this factor identified an optimal pore size of approximately 500 μm for both Diamond and Gyroid. Besides, based on such criterion, Gyroid exhibited greater applicability as bone scaffolds. In summary, this study provides comprehensive assessment of the effect of pore size and demonstrates the efficient estimation of an in-silico framework for evaluating lattice structures as medical implants, which could be applied to other lattice architectures.

Entomopathogenic fungi utilize specific secondary metabolites to defend against insect immunity, thereby enabling colonization of their specific hosts. We are particularly interested in the polyketide synthesis gene pks15, which is involved in metabolite production, and its role in fungal virulence. Targeted disruption of pks15 followed by genetic complementation with a functional copy of the gene would allow for functional characterization of this secondary metabolite biosynthesis gene. Using a Beauveria bassiana ∆pks15 mutant previously disrupted by a bialophos-resistance (bar) cassette, we report here an in-cis complementation at bar cassette using CRISPR/Cas9 gene editing. A bar-specific short guide RNA was used to target and cause a double-strand break in bar, and a donor DNA carrying a wild-type copy of pks15 was co-transformed with the guide RNA. Isolate G6 of ∆pks15 complemented with pks15 was obtained and verified by PCR, Southern analyses and DNA sequencing. Compared to ∆pks15 which showed a marked reduction in sporulation and insect virulence, the complementation in G6 restored with insect virulence, sporulation and conidial germination to wild-type levels. Atomic force and scanning electron microscopy revealed that G6 and wild-type conidial wall surfaces possessed the characteristic rodlet bundles and rough surface while ∆pks15 walls lacked the bundles and were relatively smoother. Conidia of ∆pks15 were larger and more elongated than that of G6 and the wild type, indicating changes in their cell wall organization. Our data indicate that PKS15 and its metabolite are likely not only important for fungal virulence and asexual reproduction, but also cell wall formation.

Generally, the injection method is recommended as the best efficient method for vaccine applications in fish. However, labor-intensive and difficult injection for certain fish sizes is always considered as a limitation to aquatic animals. To demonstrate the effectiveness of a novel oral delivery system for the piscine vaccine with nano-delivery made from nano clay, halloysite nanotubes (HNTs) and their modified forms were loaded with killed vaccines, and we determined the ability of the system in releasing vaccines in a mimic digestive system. The efficaciousness of the oral piscine vaccine nano-delivery system was evaluated for its level of antibody production and for the level of disease prevention in tilapia. Herein, unmodified HNTs (H) and modified HNTs [HNT-Chitosan (HC), HNT-APTES (HA) and HNT-APTES-Chitosan (HAC)] successfully harbored streptococcal bivalent vaccine with inactivated S. agalactiae, designated as HF, HAF, HCF and HACF. The releasing of the loading antigens in the mimic digestive tract demonstrated a diverse pattern of protein releasing depending on the types of HNTs. Remarkably, HCF could properly release loading antigens with relevance to the increasing pH buffer. The oral vaccines revealed the greatest elevation of specific antibodies to S. agalactiae serotype Ia in HCF orally administered fish and to some extent in serotype III. The efficacy of streptococcal disease protection was determined by continually feeding with HF-, HAF-, HCF- and HACF-coated feed pellets for 7 days in the 1st and 3rd week. HCF showed significant RPS (75.00 ± 10.83%) among the other tested groups. Interestingly, the HCF-treated group exhibited noticeable efficacy similar to the bivalent-vaccine-injected group (RPS 81.25 ± 0.00%). This novel nano-delivery system for the fish vaccine was successfully developed and exhibited appropriated immune stimulation and promised disease prevention through oral administration. This delivery system can greatly support animals’ immune stimulation, which conquers the limitation in vaccine applications in aquaculture systems. Moreover, this delivery system can be applied to carrying diverse types of biologics, including DNA, RNA and subunit protein vaccines.

Polymer composite scaffolds hold promise in bone tissue engineering due to their biocompatibility, mechanical properties, and reproducibility. Among these materials, polylactic acid (PLA), a biodegradable plastics has gained attention for its processability characteristics. However, a deeper understanding of how PLA scaffold surface properties influence cell behavior is enssential for advancing its applications. In this study, 3D-printed PLA scaffolds containing hydroxyapatite (HA) were analyzed using atomic force microscopy and nanomechanical mapping. The addition of HA significantly increased key surface properties compared to unmodified PLA scaffols. Notably, the HA-modified scaffold demonstrated Gaussian distribution of stiffness and adhesive forces, in contrast to the bimodal properties observed in the unmodified PLA scaffolds. Human adipose-derived mesenchymal stem cell (hADMSC) seeded on the 3D-printed PLA scaffolds blended with 10% HA (P10) exhibited strong attachment. After four weeks, osteogenic differentiation of hADMSCs was detected, with calcium deposition reaching 6.76% ± 0.12. These results suggest that specific ranges of stiffness and adhesive forces of the composite scaffold can support cell attachement, and mineralization. The study highlights that tailoring suface properties of composite scaffolds is crucial for modulating cellular interactions, thus advancing the development of effective bone replacement materials.

Bacteriophages offer a sustainable alternative for controlling crop disease. However, the lack of knowledge on phage infection mechanisms makes phage-based biological control varying and ineffective. In this work, we interrogated the temperature dependence of the infection and thermo-responsive behavior of the C22 phage. This soilborne podovirus is capable of lysing Ralstonia solanacearum, causing bacterial wilt disease. We revealed that the C22 phage could better infect the pathogenic host cell when incubated at low temperatures (25, 30 °C) than at high temperatures (35, 40 °C). Measurement of the C22 phage stiffness revealed that the phage stiffness at low temperatures was 2–3 times larger than at high temperatures. In addition, the imaging results showed that more C22 phage particles were attached to the cell surface at low temperatures than at high temperatures, associating the phage stiffness and the phage attachment. The result suggests that the structure and stiffness modulation in response to temperature change improve infection, providing mechanistic insight into the C22 phage lytic cycle. Our study signifies the need to understand phage responses to the fluctuating environment for effective phage-based biocontrol implementation.

Bacteriophages have potential for use as biological control agents (biocontrols) of pathogenic bacteria, but their low stability is limiting for their utilization as biocontrols. Understanding of the conditions conducive to storage of phages in which infectivity is maintained over long periods will be useful for their application as biocontrols. We employed a nanomechanical approach to study how external environmental factors affect surface properties and infectivity of the podovirus C22 phage, a candidate for biocontrol of Ralstonia solanacearum, the agent of bacterial wilt in crops. We performed atomic force microscopy (AFM)-based nano-indentation on the C22 phage in buffers with varying pH and ionic strength. The infectivity data from plaque assay in the same conditions revealed that an intermediate range of stiffness was associated with phage titer that remained consistently high, even after prolonged storage up to 182 days. The data are consistent with the model that C22 phage must adopt a metastable state for maximal infectivity, and external factors that alter the stiffness of the phage capsid lead to perturbation of this infective state.