A review of plant-based expression systems as a platform for single-domain recombinant antibody production.

Monoclonal antibodies have contributed to improving the treatment of several diseases. However, limitations related to pharmacokinetic parameters and production costs have instigated the search for alternative products. Camelids produce functional immunoglobulins G devoid of light chains and CH1 domains, in which the antigenic recognition site is formed by a single domain called VHH or nanobody. VHHs' small size and similarity to the human VH domain contribute to high tissue penetration and low immunogenicity. In addition, VHHs provide superior antigen recognition compared to human antibodies, better solubility and stability. Due to these characteristics and the possibility of obtaining gene-encoding VHHs, applications of this biological tool, whether as a monomer or in related recombinant constructs, have been reported. To ensure antibody efficacy and cost-effectiveness, strategies for their expression, either using prokaryotic or eukaryotic systems, have been utilized. Plant-based expression systems are useful for VHH related constructs that require post-translational modifications. This system has exhibited versatility, low-cost upstream production, and safety. This article presents the main advances associated to the heterologous expression of VHHs in plant systems. Besides, we show insights related to the use of VHHs as a strategy for plant pathogen control and a tool for genomic manipulation in plant systems.

In vitro inhibitory activity of terpenic derivatives against clinical and environmental strains of the Sporothrix schenkii complex.

Sporotrichosis is a subacute or chronic subcutaneous infection, caused by the fungus Sporothrix schenkii complex, occurring in human and animal tissues. Potassium iodide and itraconazole have been used as effective therapy for first-choice treatment, while amphotericin B may be indicated for disseminated infection. However, the adverse effects of potassium iodide and amphotericin B or the long duration of therapy with itraconazole often weigh against their use, leading to the search for alternatives for the treatment of severe infections. Terpinen-4-ol and farnesol are components of essential oils present in many plant species and have been described to have antifungal activity against microorganisms. In this study, 40 strains of Sporothrix spp. were tested for the susceptibility to terpinen-4-ol and farnesol. Changes in cytoplasmic membrane permeability were also investigated. Terpenes inhibited all Sporothrix strains with MIC values ranging from 87.9 to 1,429.8 µg/ml for terpinen-4-ol and from 0.003 to 0.222 µg/ml for farnesol. The MFC values ranged from 177.8 to 5,722.6 µg/ml and from 0.027 to 0.88 µg/ml, respectively, for terpinen-4-ol and farnesol. Farnesol was the most active compound for the Sporothrix strains. Significant loss of 260 and 280 nm-absorbing material did not occur after treatment with concentrations equivalent to the MIC and sub-MIC of the tested terpenes, when compared to corresponding untreated samples. The failure of terpenes to lyse Sporothrix cells suggests that their primary mechanism of action is not by causing irreversible cell membrane damage. Thus, new studies are needed to better understand the mechanisms involved in the antifungal activity.

In vitro inhibitory effect of miltefosine against strains of Histoplasma capsulatum var. capsulatum and Sporothrix spp.

Miltefosine (MIL), originally developed for use in cancer chemotherapy, has been shown to have important antifungal activity against several pathogenic fungi. Our aim in this study was to determine the in vitro activity of MIL against the dimorphic fungi Histoplasma capsulatum and Sporothrix spp. This was done using the broth microdilution method. MIL had an in vitro inhibitory effect against all strains of H. capsulatum var. capsulatum and Sporothrix spp. analyzed. The minimal inhibitory concentrations (MIC) varied from 0.25 µg/ml to 2 µg/ml for H. capsulatum var. capsulatum in the filamentous phase and from 0.125 µg/ml to 1 µg/ml in the yeast phase. The MIC interval for Sporothrix spp. in the filamentous phase was 0.25-2 µg/ml. The minimal fungicidal concentrations (MFCs) were ≤4 µg/ml for isolates of both analyzed species. This study demonstrates that MIL has an antifungal effect in vitro against two potentially pathogenic fungi and that more studies should be performed in order to evaluate its applicability in vivo.