Parallel to the growing use of kratom, there is a wealth of evidence from self-report, preclinical, and early clinical studies on therapeutic benefits of its alkaloids in particular for treating pain, managing substance use disorder, and coping with emotional or mental health conditions. On the other hand, there are also reports on potential health risks concerning kratom use. These two aspects are often discussed in reviews on kratom. Here, we aim to highlight specific areas that are of importance to give insights into the mechanistic of kratom alkaloids pharmacological actions. This includes their interactions with drug-metabolizing enzymes and predictions of clinical drug-drug interactions, receptor-binding properties, interactions with cellular barriers in regards to barrier permeability, involvement of membrane transporters, and alteration of barrier function when exposed to the alkaloids.

Enterovirus A71 (EV-A71) causes hand, foot and mouth disease epidemics with neurological complications and fatalities. However, the neuropathogenesis of EV-A71 remains poorly understood. In mice, adaptation and virulence determinants have been mapped to mutations at VP2-149, VP1-145 and VP1-244. We investigate how these amino acids alter heparin-binding phenotype and shapes EV-A71 virulence in one-day old mice. We constructed six viruses with varying residues at VP1-98, VP1-145 (which are both heparin-binding determinants) and VP2-149 (based on the wild type 149K/98E/145Q, termed KEQ) to generate KKQ, KKE, KEE, IEE and IEQ variants. We demonstrated that the weak heparin-binder IEE was highly lethal in mice. The initially strong heparin-binding IEQ variant acquired an additional mutation VP1-K244E, which confers weak heparin-binding phenotype resulting in elevated viremia and increased brain virus antigens in mice, with subsequent high virulence. IEE and IEQ-244E variants inoculated into mice disseminated efficiently and displayed high viremia. Increasing polymerase fidelity and impairing recombination of IEQ attenuated the virulence, suggesting the importance of population diversity in EV-A71 pathogenesis in vivo. Combining in silico docking and deep sequencing approaches, we inferred that virus population diversity is shaped by electrostatic interactions at the five-fold axis of the virus surface. Electrostatic surface charges facilitate virus adaptation by generating poor heparin-binding variants for better in vivo dissemination in mice, likely due to reduced adsorption to heparin-rich peripheral tissues, which ultimately results in increased neurovirulence. The dynamic switching between heparin-binding and weak heparin-binding phenotype in vivo explained the neurovirulence of EV-A71. Enterovirus A71 (EV-A71) is the primary cause of hand, foot and mouth disease, and it can also infect the central nervous system and cause fatal outbreaks in young children. EV-A71 pathogenesis remains elusive. In this study, we demonstrated that EV-A71 variants with strong affinity to heparan sulfate (heparin) have a growth advantage in cell culture, but are disadvantaged in vivo. When inoculated into one-day old mice, strong heparin-binding virus variants are more likely to be adsorbed to peripheral tissues, resulting in impaired ability to disseminate, and are cleared from the bloodstream rapidly. The lower viremia level resulted in no neuroinvasion. In contrast, weak heparin-binding variants show greater levels of viremia, dissemination and subsequent neurovirulence in mice. We also provide evidence that the EV-A71 heparin-binding pattern is mediated by electrostatic surface charges on the virus capsid surface. In mice, EV-A71 undergoes adaptive mutation to acquire greater negative surface charges, thus generating new virulent variants with weak heparin-binding ability which allows greater viral spread. Our study underlines the importance of electrostatic surface charges in shaping EV-A71 virulence.