Genomic epidemiology of SARS-CoV-2 in large university hospital cohort: the UnCoVER-Brazil project.

This work aimed to study the role of different SARS-CoV-2 lineages in the epidemiology of multiple waves of the COVID-19 pandemic in Ribeirão Preto (São Paulo state), with comparison within Brazil and globally. Viral genomic sequencing was combined with clinical and sociodemographic information of 2,379 subjects at a large Brazilian hospital. On the whole 2,395 complete SARS-CoV-2 genomes were obtained from April 2020 to January 2022. We report variants of concern (VOC) and interest (VOI) dynamics and the role of Brazilian lineages. We identified three World Health Organization VOCs (Gamma, Delta, Omicron) and one VOI (Zeta), which caused distinct waves in this cohort. We also identified 47 distinct Pango lineages. Consistent with the high prevalence of Gamma in Brazil, Pango lineage P.1 dominated infections in this cohort for half of 2021. Each wave of infection largely consisted of a single variant group, with each new group quickly and completely rising to dominance. Despite increasing vaccination in Brazil starting in 2021, this pattern was observed throughout the study and is consistent with the hypothesis that herd immunity tends to be SARS-CoV-2 variant-specific and does not broadly protect against COVID-19.

A Simple and Cost-Efficient Platform for a Novel Porcine Circovirus Type 2d (PCV2d) Vaccine Manufacturing.

Porcine circovirus type 2d (PCV2d) is becoming the predominant PCV genotype and considerably affects the global pig industry. Nevertheless, currently, no commercial PCV2d vaccine is available. Preventing and controlling the disease caused by PCV2d is therefore based on other genotype-based vaccines. However, their production platforms are laborious, limited in expression level, and relatively expensive for veterinary applications. To address these challenges, we have developed a simple and cost-efficient platform for a novel PCV2d vaccine production process, using fed-batch E. coli fermentation followed by cell disruption and filtration, and a single purification step via cation exchange chromatography. The process was developed at bench scale and then pilot scale, where the PCV2d subunit protein yield was approximately 0.93 g/L fermentation volume in a short production time. Moreover, we have successfully implemented this production process at two different sites, in Southeast Asia and Europe. This demonstrates transferability and the high potential for successful industrial production.

Transmission from vaccinated individuals in a large SARS-CoV-2 Delta variant outbreak.

An outbreak of over 1,000 COVID-19 cases in Provincetown, Massachusetts (MA), in July 2021-the first large outbreak mostly in vaccinated individuals in the US-prompted a comprehensive public health response, motivating changes to national masking recommendations and raising questions about infection and transmission among vaccinated individuals. To address these questions, we combined viral genomic and epidemiological data from 467 individuals, including 40% of outbreak-associated cases. The Delta variant accounted for 99% of cases in this dataset; it was introduced from at least 40 sources, but 83% of cases derived from a single source, likely through transmission across multiple settings over a short time rather than a single event. Genomic and epidemiological data supported multiple transmissions of Delta from and between fully vaccinated individuals. However, despite its magnitude, the outbreak had limited onward impact in MA and the US overall, likely due to high vaccination rates and a robust public health response.

Evidence of transmission from fully vaccinated individuals in a large outbreak of the SARS-CoV-2 Delta variant in Provincetown, Massachusetts.

Multiple summer events, including large indoor gatherings, in Provincetown, Massachusetts (MA), in July 2021 contributed to an outbreak of over one thousand COVID-19 cases among residents and visitors. Most cases were fully vaccinated, many of whom were also symptomatic, prompting a comprehensive public health response, motivating changes to national masking recommendations, and raising questions about infection and transmission among vaccinated individuals. To characterize the outbreak and the viral population underlying it, we combined genomic and epidemiological data from 467 individuals, including 40% of known outbreak-associated cases. The Delta variant accounted for 99% of sequenced outbreak-associated cases. Phylogenetic analysis suggests over 40 sources of Delta in the dataset, with one responsible for a single cluster containing 83% of outbreak-associated genomes. This cluster was likely not the result of extensive spread at a single site, but rather transmission from a common source across multiple settings over a short time. Genomic and epidemiological data combined provide strong support for 25 transmission events from, including many between, fully vaccinated individuals; genomic data alone provides evidence for an additional 64. Together, genomic epidemiology provides a high-resolution picture of the Provincetown outbreak, revealing multiple cases of transmission of Delta from fully vaccinated individuals. However, despite its magnitude, the outbreak was restricted in its onward impact in MA and the US, likely due to high vaccination rates and a robust public health response.

Testicular pain following initiation of elexacaftor/tezacaftor/ivacaftor in males with cystic fibrosis.

Elexacaftor/tezacaftor/ivacaftor (Trikafta) was approved by the Food and Drug Administration in October 2019 for treatment of cystic fibrosis (CF) in patients 12 years and older with at least one F508del mutation in the CFTR protein. There were no documented reports of testicular pain during clinical trials. In this case series, we discuss 7 males between 17 and 39 years of age who reported testicular pain or discomfort within the first two weeks of starting therapy. The precise mechanism of this side effect is unknown, but it may be related to restoration of CFTR function in the male reproductive tract. All patients experienced resolution of this side effect within a week after onset, regardless of the management, except for one. Further research is needed to determine short- and long-term impact of this drug on male fertility. Until more data is available, the authors recommend counseling patients on contraceptive options.

Trypanosoma cruzi infections and associated pathology in urban-dwelling Virginia opossums (Didelphis virginiana).

Trypanosoma cruzi, a zoonotic protozoan parasite, infects a wide range of mammals. The southern United States has endemic sylvatic transmission cycles maintained by several species of wildlife and domestic dogs. We hypothesized that urban-dwelling opossums (Didelphis virginiana) in South Texas are infected with T. cruzi, and that tissue pathology would be associated with infection. In 2017, we collected blood, heart tissue and anal gland secretions from 100 wild opossums across three seasons that were trapped by animal control in South Texas. In addition, anal gland tissue and intercostal muscle were collected from 43 of the 100 opossums for which time allowed the extra tissue collection. All blood, tissue, and secretion samples were screened for T. cruzi DNA using qPCR with confirmation of positive status achieved through one or more additional PCR assays, including a qPCR to determine the parasite discrete typing unit (DTU). T. cruzi DNA was detected in at least one tissue of 15% of the opossums sampled: blood clot (9%), heart tissue (10%), anal gland secretions (12%), intercostal muscle (16.3%), and anal gland tissue (11.6%). Infection was detected in two or more different tissue types in nine of the opossums. The 35 tissues for which parasite DTU was determined were exclusively 'Tcl'- a DTU previously associated with locally-acquired human disease in the United States. T. cruzi-positive opossums were nearly 14 times more likely to exhibit significant heart lesions on histopathology (lympoplasmacytic inflammation±fibrosis) when compared to negative opossums (OR = 13.56, CI = 1.23-751.28, p-value = 0.03). Three triatomines were opportunistically collected from the study site, of which two were infected (66.7%), and bloodmeal analysis revealed canine, opossum, and human bloodmeals. Given the presence of parasite in opossum blood, unique potential for shedding of parasite in anal glad secretions, and evidence of vectors feeding on opossums, it is likely that opossums serve as wild reservoirs around urban dwellings in South Texas.

Prevalence of Trypanosoma cruzi infection and associated histologic findings in domestic cats (Felis catus).

Trypanosoma cruzi is a zoonotic protozoan parasite transmitted by triatomines that infects a wide range of mammals. South Texas is a hotspot for triatomines, T. cruzi-infected dogs and wildlife, and local transmission to humans also occurs. However, little is known about the infection of domestic cats (Felis catus) in the United States. Given the role cats play in the ecology of T. cruzi in Mexico and South America, we hypothesized that T. cruzi infection occurs in cats from south Texas, sometimes associated with cardiac pathology. In 2017, 167 euthanized cats from a south Texas shelter were sampled across winter, spring, and summer. We collected whole blood and hearts from all cats, with additional tissues from a subset. Serum samples were screened for T. cruzi antibodies using two independent rapid immunochromatographic tests and an indirect fluorescent antibody test. Cats were considered seropositive if they were positive on at least two independent serological tests. Blood clot, heart tissue and other tissues were subjected to qPCR for parasite detection and discrete typing unit (DTU) determination. Tissues from selected seropositive or PCR-positive animals and a subset of negative animals were processed routinely for histopathology and examined by a board-certified pathologist. A total of 19 cats (11.4%) were seropositive and three cats (1.8%) - one of which was seropositive - had one or more PCR-positive tissues. Infected tissues included heart, bicep femoris muscle, sciatic nerve, esophagus, and mesentery. Genotyping of the parastite to the level of DTU showed that exclusively DTU TcI was present, despite past studies showing both TcI and TcIV in vectors of the region. Eight of 19 (42.1%) seropositive cats exhibited lymphoplasmacytic inflammation, sometimes with fibrosis, in cardiac tissue compared to 28.6% of 28 seronegative cats (P = 0.10). Domestic cats are affected hosts in the eco-epidemiology of Chagas disease. Future prospective studies are needed to understand disease progression. Veterinarians in the southern United States should consider T. cruzi in their index of suspicion in cats with exposure to vectors and undetermined cardiac abnormalities.

Hybrid nanocarriers incorporating mechanistically distinct drugs for lymphatic CD4+ T cell activation and HIV-1 latency reversal.

A proposed strategy to cure HIV uses latency-reversing agents (LRAs) to reactivate latent proviruses for purging HIV reservoirs. A variety of LRAs have been identified, but none has yet proven effective in reducing the reservoir size in vivo. Nanocarriers could address some major challenges by improving drug solubility and safety, providing sustained drug release, and simultaneously delivering multiple drugs to target tissues and cells. Here, we formulated hybrid nanocarriers that incorporate physicochemically diverse LRAs and target lymphatic CD4+ T cells. We identified one LRA combination that displayed synergistic latency reversal and low cytotoxicity in a cell model of HIV and in CD4+ T cells from virologically suppressed patients. Furthermore, our targeted nanocarriers selectively activated CD4+ T cells in nonhuman primate peripheral blood mononuclear cells as well as in murine lymph nodes, and substantially reduced local toxicity. This nanocarrier platform may enable new solutions for delivering anti-HIV agents for an HIV cure.

TrkB expression and phospho-ERK activation by brain-derived neurotrophic factor in rat spinothalamic tract neurons.

Brain-derived neurotrophic factor (BDNF) is a neurotrophin implicated in the phenomena of synaptic plasticity in the adult. It is found in terminals of nociceptive primary afferents. Following a pain-related stimulus, it is released in the spinal cord, where it activates its high-affinity receptor TrkB, leading to the phosphorylation of the mitogen-activated protein kinase (MAPK) extracellular signal-regulated kinase (ERK). A large body of evidence suggests that BDNF has a positive neuromodulatory effect on glutamate transmission in the spinal cord. However, none of these studies examined anatomically whether projection neurons known to be involved in transmission of nociceptive inputs express BDNF's receptor. Because the spinothalamic tract (STT) is a well-characterized pathway for its role in the transfer and integration of sensory and nociceptive informations, this study in rats aimed to 1) determine whether neurons of the STT pathway express the TrkB receptor, 2) establish the rostrocaudal and laminar distribution of STT-TrkB neurons in the whole spinal cord, and 3) test the potential functionality of TrkB expression in these cells by investigating the ability of BDNF to activate the MAP kinase ERK. Using tract tracing coupled to immunofluorescent labeling for TrkB, we observed that in all levels of the spinal cord most STT neurons were immunoreactive for TrkB. Furthermore, microinjections of BDNF into the spinal cord or release of endogenous BDNF by intraplantar injection of capsaicin activated ERK phosphorylation in TrkB-containing STT neurons. These data suggest an important role for BDNF in nociception as an activator of spinothalamic projection neurons.

Brain-derived neurotrophic factor induces NMDA receptor subunit one phosphorylation via ERK and PKC in the rat spinal cord.

Brain-derived neurotrophic factor (BDNF) is involved in the modulation of synaptic transmission in the spinal cord, and several circumstantial lines of evidence suggest that it has the ability to modulate the activity of the NMDA receptor. Here we dissect the signalling mechanisms by which BDNF exerts its neuromodulatory role on the NMDA receptor subunit 1 (NR1). Using a preparation of adult isolated dorsal horn with dorsal roots attached, we found that electrical stimulation of roots induced a concomitant release of BDNF and an increased phosphorylation of NR1, which was partly prevented by the BDNF sequestering molecule, TrkB-IgG. Using a second approach in vitro, we confirmed that both exogenous glutamate and BDNF (but not other neurotrophins) were able to induce NR1 phosphorylation, in particular at residue Ser-897. NR1 phosphorylation induced by BDNF was blocked by a TrkB inhibitor, an ERK inhibitor and a PKC inhibitor but not a PKA inhibitor. Activation of PKC using exogenous PMA also led to NR1 phosphorylation. Together these data suggest that BDNF modulates the activity of the receptor by phosphorylation via the kinases ERK and PKC.

Brain-derived neurotrophic factor induces NMDA receptor 1 phosphorylation in rat spinal cord.

The NMDA receptor and the neurotrophin brain-derived neurotrophic factor, BDNF, are involved in central sensitisation and synaptic plasticity in the spinal cord. Recent evidence suggests that BDNF modulates NMDA responses, by a yet unknown mechanism. The phosphorylation of the NMDA subunit 1 following BDNF exposure has been investigated in the neonatal rat spinal cord. Western blotting revealed that BDNF administered to isolated cords for 20 minutes significantly elevated phospho-NR1 levels. Immunohistochemical analysis localised this increase to functionally appropriate regions of the dorsal horn. Pre-incubation with trkB-IgG (BDNF sequestering molecule) abolished this NR1 phosphorylation. We suggest that BDNF modulates synaptic activity within the spinal cord via NR1 phosphorylation.

Expression of gp130 and leukaemia inhibitory factor receptor subunits in adult rat sensory neurones: regulation by nerve injury.

Members of the interleukin-6 (IL-6) family of cytokines have been implicated as major mediators of the response of the adult nervous system to injury. The basis for an interaction of IL-6 cytokines with adult sensory neurones has been established by analysing the levels and distribution of the two signal-transducing receptor subunits, glycoprotein 130 (gp130) and leukaemia inhibitory factor receptor (LIFR), in the dorsal root ganglion (DRG) of male adult rats before and following nerve injury. All sensory neurones express gp130-immunoreactivity (IR) in the cytoplasm and on the plasma membrane. Levels of gp130 and its intracellular distribution remained unchanged up to 14 days following sciatic nerve axotomy. LIFR-IR was largely absent from the cytoplasm and plasma membrane of sensory neurones, but confined almost exclusively to the nuclear compartment. However, following axotomy, punctate cytoplasmic LIFR-IR was detected which persisted up to 28 days following axotomy. The expression of cytoplasmic LIFR 2 days post-axotomy was proportionally greater in a subset of small diameter sensory neurones which expressed either the sensory neuropeptide CGRP or the cell surface marker isolectin B4. The coexpression of gp130 and LIFR in the same intracellular compartment following axotomy conveys potential responsiveness of injured sensory neurones to members of the IL-6 family of cytokines.