Gregory P. Contreras

Despite advances in drug discovery, effective delivery and precise imaging still pose significant obstacles. Scientists have turned to zero-dimensional nanomaterials like carbon dots (CDs) to enable precision drug delivery. Due to their multidisciplinary appeal, there is extensive research on both the physical properties and biological applications of CDs, yet the fields rarely complement each other. This review applies a quantum-mechanistic framework to better understand CDs and unify the relationship between electronic structure, photophysical behavior, and theranostic performance. We first highlight the theranostic advantages of CDs over quantum dots, then discuss the quantum mechanisms behind a CD’s imaging capabilities, including surface states, excitation-dependent photoluminescence, and the electronic origins of photostability. Through this physical lens, we then evaluate the effect of different precursors and doping strategies during synthesis on electronic structure and how dots are engineered to load, release, and track therapeutic cargo. Photodynamic therapy and surface functionalization strategies are also evaluated. Finally, we address the significance of renal clearance and translational bottlenecks such as heterogeneity.

Viral recombination can rapidly generate new variants and influence outbreak severity. Currently, a method to track viral recombination events at high genomic resolution during natural transmission remains unavailable. Here we describe a novel linkage disequilibrium method and the first report of viral recombination events over time by using single nucleotide polymorphisms (SNPs) as natural genetic markers. Combined with phylogenetic data, we analyzed mpox virus (MPXV) Ib genome during 2023-2025 outbreak and found that 71 recombination events among Ib genomes, yielding a recombination frequency (Rf) and rate of 0.16%/kbp and 0.11%/kbp/year, respectively. Phylogenetic time analysis also shows that recombination began as early as December 23, 2023. Rf of MPXV Ib is 8.8-fold lower compared to vaccinia virus (1.5%/kbp) in vitro . Recombination and linkage hotspots are enriched at both inverted terminal repeat and variable regions of MPXV Ib genome. Our data not only reports the first Rf and rate of natural human transmission in the poxvirus family but also provides a powerful genomic surveillance tool for better understanding transmission and evolution of any viral outbreak globally.

HKU5-CoV-2 is a new bat-infecting coronavirus phylogenetically related to MERS-CoV. It has recently been confirmed that HKU5-CoV-2 can enter human cells and organoids in vitro via the ACE2 receptor, raising concerns about its pandemic potential due to zoonotic spillover. Whether recombination has an influence on HKU5-CoV-2 is completely unclear to date. Here we report the first evidence of HKU5-CoV-2 viral recombination, in association with mutations at the receptor binding domain (RBD) and S1/S2 furin cleavage site (FCS) of the spike protein. Using linkage disequilibrium and haploblock analysis, we identified that 167 recombination hotspots and 27 haploblocks. SNP23016/23043/23064/23156/23193/23285 at the RBD and SNP23833/23847 at the FCS are recombinant hotspots. Our results suggest that recombination may lead to the substitution at RBD residue 498 (Thr498Val/Val498Thr, Thr498Ile/Ile498Thr), which Thr498 directly contacts the ACE2 receptor. Recombination also causes Ser723 deletion/insertion and Ser729Ala substitution at the FCS. These mutations could affect host tropism and change furin cleavage activity. Our results indicate that recombination has played a critical role in HKU5-CoV-2 evolution and infectivity.

The World Health Organization recently declared the 2024 mpox virus (MPXV Ib) outbreak a public health emergency of international concern. We report that in 2023-2024, MPXV clade Ib genomes are diverging at a faster rate than clade IIb (2022), primarily due to an unusually high incidence of recombination. Phylogenetic analysis revealed that Ib strains have diverged into four lineages, and they have evolved into 14 subgroups based on nine tandem repeat (TR) polymorphisms. These findings confirms that TRs in MPXV Ib are mutating at a significantly higher frequency compared to the 2022 outbreak (clade IIb, 11 subgroups). Linkage disequilibrium analysis also identified 10 recombination clusters among all 4 lineages, with recombination incidence in Ib being twice as high as in IIb. This suggests that a higher rate of superinfection is contributing to ongoing recombination among populations infected with clade Ib. Prompt action is necessary to prevent the emergence of more lethal mpox strains.

Many integrative filamentous phages not only lack Ff coliphage homologues essential for assembly but also have distinct packaging signals (PS). Their encapsidation remains completely uncharacterized to date. Here we report the first evidence of a PS-dependent checkpoint for integrative filamentous phage assembly. Suppressor screening of PS-deficient phages identified an unknown protein, PSB15 (PS-binding 15 kDa), crucial for encapsidation. The WAGFXF motif of the PSB15 N-terminus directly binds to PS DNA with conformational change, while suppressor mutations relieve DNA binding specificity constraints to rescue assembly arrest. PSB15 interacts with phospholipid cardiolipin via its basic helix and C-terminus, and recruits PS DNA to the inner membrane (IM). The PSB15-PS complex is released from the IM by interaction between its hydrophobic linker and thioredoxin (Trx), a host protein that is required for Ff assembly but whose mechanisms are still unclear. Live cell imaging shows that thioredoxin and DNA binding regulate the dwelling time of PSB15 at cell poles, suggesting that they both facilitate the dissociation of PSB15 from the IM. Loss of PSB15 or its PS-binding and IM-targeting/dissociation activity compromised virus egress, indicating that the PS/PSB15/Trx complex establishes a regulatory phage assembly checkpoint critical for integrative phage infection and life cycles.

Many integrative filamentous phages not only lack Ff coliphage homologues essential for assembly but also have distinct packaging signals (PS). Their encapsidation remains completely uncharacterized to date. Here we report the first evidence of a PS-dependent checkpoint for integrative filamentous phage assembly. Suppressor screening of PS-deficient phages identified an unknown protein, PSB15 ( PS - b inding 15 kDa), crucial for encapsidation. The WAGFXF motif of the PSB15 N-terminus directly binds to PS DNA with conformational change, while suppressor mutations relieve DNA binding specificity constraints to rescue assembly arrest. PSB15 interacts with phospholipid cardiolipin via its basic helix and C-terminus, and recruits PS DNA to the inner membrane (IM). The PSB15-PS complex is released from the IM by interaction between its hydrophobic linker and thioredoxin (Trx), a host protein that is required for Ff assembly but whose mechanisms are still unclear. Live cell imaging shows that thioredoxin and DNA binding regulate the dwelling time of PSB15 at cell poles, suggesting that they both facilitate the dissociation of PSB15 from the IM. Loss of PSB15 or its PS-binding and IM-targeting/dissociation activity compromised virus egress, indicating that the PS/PSB15/Trx complex establishes a regulatory phage assembly checkpoint critical for integrative phage infection and life cycles.

Unlike Ff, the packaging signal (PS) and the mechanism of integrative filamentous phage assembly remain largely unknown. Here we revived two Inoviridae prophage sequences, ϕLf2 and ϕLf-UK, as infectious virions lysogenize black rot pathogen Xanthomonas campestris pv. campestris . ϕLf2 and ϕLf-UK genomes consist of 6,363 and 6,062 nucleotides and share 85.8% and 98.7% identity with ϕLf, respectively. To explore their assembly, we first identified 20-26– nucleotide long PS sequences of 10 Xanthomonas phages. These PS consist of a DNA hairpin with the consensus GGX(A/-)CCG(C/T)G sequence in the stem and C/T nucleotides in the loop, both of which are conserved and essential for PS activity. In contrast to Ff, the 5’ to 3‘ orientation of PS sequence is not conserved or critical for their competence. This is the first report to offer insights into the structure and function of integrative phage PS, revealing the diversity of filamentous phage encapsidation.

Here we report the first evidence of recombination of monkeypox genome in natural transmission by analyzing six tandem repeats (TRs) among 415 sequences collected between January to July 2022. The 2022 monkeypox viral population has diverged into 11 subgroups based on various TRs and their copy numbers. Here we identify 8 new recombinants (six from Slovenia, one from Australia, one from Italy). Our results indicate that the monkeypox genome is evolving and expanding quickly during the 2022 pandemic (Zeng’s E = -1.65, Achaz’s Y=-2.52, p< 0.001). We conclude that, In combination with genomic surveillance, TR analysis is a useful tool to monitor and track phylogenetic dynamics of monkeypox transmission.

On 26 November 2021, the World Health Organization designated the SARS-CoV-2 variant B.1.1.529, Omicron, a variant of concern. However, the phylogenetic and evolutionary dynamics of this variant remain unclear. An analysis of the 131 Omicron variant sequences from November 9 to November 28, 2021 reveals that variants have diverged into at least 6 major subgroups. 86.3% of the cases have an insertion at amino acid 214 (INS214EPE) of the spike protein. Neutrality analysis of DH (−2.814, p E (0.0583, p =1.0) tests suggested that directional selection was the major driving force of Omicron variant evolution. The synonymous ( D _syn ) and nonsynonymous ( D _nonsyn ) polymorphisms of the Omicron variant spike gene were estimated with Tajima’s D statistic to eliminate homogenous effects. Both D ratio ( D _nonsyn / D _syn , 1.57) and Δ D ( D _syn - D _nonsyn , 0.63) indicate that purifying selection operates at present. The low nucleotide diversity (0.00008) and Tajima D value (−2.709, p D test score, with a threshold value as −2.50, as an accurate predictor of new COVID-19 outbreaks.

Public distrust has undermined COVID-19 vaccine acceptance and has become a major public health issue in the battle against SARS-CoV-2 transmission globally. Here we present the first evidence that the vaccination coverage rate is inversely correlated to the mutation frequency of the SARS-CoV-2 delta variant in 16 countries (R²=0.878), strongly indicating that full vaccination against COVID-19 is critical to suppress emergent mutations. We also present a promising tool to forecast new COVID-19 outbreaks. The Tajima’s D test, an evolutionary algorithm, with a threshold value of -2.50 is shown to be an accurate predictor of new outbreaks. We recommend that universal vaccination, as well as mitigation strategies, and genomic surveillance continue to be employed to prevent further viral transmission.