Attraction and mating between male and female animals depend on effective communication between conspecifics. However, in mosquitoes, we have only a rudimentary understanding of the sensory cues and receptors critical for the communication that is essential for reproductive behavior. While it is known that male Aedes aegypti use sound to help them identify females, it is not unclear whether sound detection is absolutely required since other cues such as vision may also participate in mating behavior. To determine the effect of eliminating hearing on mating success, we knocked out the Ae. aegypti TRPVa channel, which is a protein expressed in chordotonal neurons in the Johnston’s organ (JO) that respond to sound-induced movements in the antenna. Loss of trpVa eradicated sound-induced responses from the JO, thereby abolishing hearing. Strikingly, mutation of trpVa eliminated mating behavior in males. In contrast, trpVa -null females mated, although this behavior was slightly delayed relative to wild-type females. Males and females produce sounds as they beat their wings at distinct frequencies during flight. Sound mimicking the female wingbeat induced flight, attraction, and copulatory-like behavior in wild-type males without females present, but not in trpVa -null males. Males are known to modulate their wingbeat frequencies before mating in the air, which is a phenomenon referred to as rapid frequency modulation (RFM). We found that RFM was absent in mosquitoes lacking TRPVa. We conclude that the requirement for trpVa and hearing for male reproductive behavior in Aedes is absolute, as mating in the deaf males is eliminated.

Central nervous system (CNS) infection with Sindbis virus (SINV), the prototypic alphavirus, results in encephalomyelitis in a well-established mouse model that is used to characterize immune factors important in controlling viral infection in neurons. We have previously shown that interferon regulatory factor (IRF) 7 is required for survival from SINV encephalomyelitis, as mice deficient in IRF7 ( Irf7 ^−/− ) develop paralysis and fatal disease within 7–8 days after infection without clearing infectious virus from the CNS. To determine the contributions of this innate immune factor in the prevention of fatal disease, we characterized the antiviral immune response to SINV infection in Irf7 ^−/− and C57BL/6J [wild-type (WT)] mice. Irf7 ^{−/ −} mice had prolonged and widespread viral infection in motor neuron-rich regions of the CNS associated with more rapid and severe immunopathology in these regions. Proportions of CD8 ⁺ T cells and inflammatory macrophages were higher in Irf7 ^−/− mice following infection, but T cells infiltrating the CNS produced fewer antiviral cytokines than in WT mice. Levels of IFN-γ and SINV-specific antibody in motor neuron-rich regions of the brain were comparable or higher than WT, indicating that IRF7 deficiency does not impair expression of these immune factors important in facilitating viral clearance. Therefore, IRF7 is required for the clearance of SINV from motor neuron-rich regions of the CNS through a mechanism that either is necessary for the neuronal response to currently identified mediators of infectious virus clearance or enables the production of additional antiviral factor(s) needed for clearance. IMPORTANCE: Viral encephalomyelitis outcome is dependent on host responses to neuronal infection. Interferon (IFN) is an important component of the innate response, and IFN regulatory factor (IRF) 7 is an inducible transcription factor for the synthesis of IFN-α. IRF7-deficient mice develop fatal paralysis after CNS infection with Sindbis virus, while wild-type mice recover. Irf7 ^−/− mice produce low levels of IFN-α but high levels of IFN-β with induction of IFN-stimulated genes, so the reason for this difference is not understood. The current study shows that Irf7 ^−/− mice developed inflammation earlier but failed to clear virus from motor neuron-rich regions of the brainstem and spinal cord. Levels of IFN-γ and virus-specific antibody were comparable, indicating that IRF7 deficiency does not impair expression of these known viral clearance factors. Therefore, IRF7 is either necessary for the neuronal response to currently identified mediators of clearance or enables the production of additional antiviral factor(s) needed for clearance.

Sindbis virus (SINV) infection of mice provides a model system for studying the pathogenesis of alphaviruses that infect the central nervous system (CNS) to cause encephalomyelitis. While studies of human viral infections typically focus on accessible cells from the blood, this compartment is rarely evaluated in mice. To bridge this gap, single-cell RNA sequencing (scRNAseq) was combined with flow cytometry to characterize the transcriptional and phenotypic changes of peripheral blood mononuclear cells (PBMCs) from SINV-infected mice. Twenty-one clusters were identified by scRNAseq at 7 days after infection, with a unique cluster and overall increase in naive B cells for infected mice. Uninfected mice had fewer immature T cells and CCR9 ⁺ CD4 T cells and a unique immature T cell cluster. Gene expression was most altered in the Ki67 ⁺ CD8 T cell cluster, with chemotaxis and proliferation-related genes upregulated. Global analysis indicated metabolic changes in myeloid cells and increased expression of Ccl5 by NK cells. Phenotypes of PBMCs and cells infiltrating the CNS were analyzed by flow cytometry over 14 days after infection. In PBMCs, CD8 and Th1 CD4 T cells increased in representation, while B cells showed a transient decrease at day 5 in total, Ly6a ⁺ , and naive cells, and an increase in activated B cells. In the brain, CD8 T cells increased for the first 7 days, while Th1 CD4 T cells and naive and Ly6a ⁺ B cells continued to accumulate for 14 days. Therefore, dynamic immune cell changes can be identified in the blood as well as the CNS during viral encephalomyelitis. IMPORTANCE: The outcome of viral encephalomyelitis is dependent on the host immune response, with clearance and resolution of infection mediated by the adaptive immune response. These processes are frequently studied in mouse models of infection, where infected tissues are examined to understand the mechanisms of clearance and recovery. However, studies of human infection typically focus on the analysis of cells from the blood, a compartment rarely examined in mice, rather than inaccessible tissue. To close this gap, we used single-cell RNA sequencing and flow cytometry to profile the transcriptomic and phenotypic changes of peripheral blood mononuclear cells (PBMCs) before and after central nervous system (CNS) infection in mice. Changes to T and B cell gene expression and cell composition occurred in PBMC and during entry into the CNS, with CCL5 being a differentially expressed chemokine. Therefore, dynamic changes occur in the blood as well as the CNS during the response of mice to virus infection, which will inform the analysis of human studies.

In the last decade, the implementation of enhanced recovery protocol for patients undergoing thoracic surgical procedures, either by thoracotomy or thoracoscopy, has gained significant recognition. Such protocols have been developed following the enhanced recovery after surgery (ERAS) guiding principles, yet have been tailored with attention to the unique nuances of thoracic surgical patients. Over the last 5 years, a body of literature has been published attesting to the success of the enhanced recovery after thoracic surgery (ERATS) protocol, with reported improvements of measurable outcome metrics. This mini-review focuses on postoperative pain control using the innovative strategy of opioid-sparing multimodal analgesics of the ERATS care pathway.

Alphaviruses are mosquito-borne viruses that can infect the central nervous system (CNS) and cause encephalomyelitis, which is a rare but dangerous complication from infection. In mice, this can be studied in a model of infection with Sindbis virus (SINV), which infects neurons and causes neurological disease. Due to the non-renewable nature of neurons, the immune response in the CNS is specialized to prevent neuronal damage or death, even if they are infected. Therefore, insights into the nuances of antiviral immunity in the CNS provide a better understanding of disease pathogenesis and mechanisms of recovery. Type I interferons (IFNs) are critically important for survival; they are an innate antiviral defense mechanism that consists mainly of IFNα and IFNβ. Although both use the same receptor, type-specific differences between IFNα and IFNβ have been described in other contexts. To this end, Ifnb^−/− mice were used to elucidate the role of IFNβ in recovery from alphavirus encephalomyelitis. IFNβ-deficient mice have intact IFNα expression and downstream signaling, but symptomatic disease occurs earlier and is more severe. This is accompanied by increased virus replication in the early stages of infection. Microgliosis is reduced in Ifnb^−/− mice compared to wildtype, but inflammatory cytokine/chemokine levels are higher and associated with alterations in monocyte and NK cell recruitment into the CNS. Ifnb^−/− mice have no deficiencies in the expression of factors known to be required for viral clearance. Therefore, IFNβ modulates the early stages of the immune response and facilitates restriction of virus replication, contributing to delayed disease onset.

Chikungunya virus, a mosquito-borne alphavirus, causes fever, rash, arthritis, and neurological disorders. Its non-structural protein 3 harbors a macrodomain, a key neurovirulence factor that removes adenosine diphosphate ribose from ADP-ribosylated substrates. Notably, chikungunya virus infection results in distinct ADP-ribosylation patterns and non-structural protein 3 macrodomain-mediated replication dynamics in astrocytes and neurons. Understanding the connection between ADP-ribosylation and the activation of innate immunity, particularly interferon release, is key to elucidating how the cellular immunological state influences ADP-ribosylation, an understudied post-translational modification during viral infection. Here, murine astrocytic (C8-D1A) and neuronal (NSC-34) cells were infected with chikungunya virus to profile transcript and protein expression of innate immune mediators and type I IFNs. The role of PARP1 in global ADP-ribosylation patterns was assessed using PARP-specific inhibitors and genetic depletion approaches. Our investigations revealed that neuronal chikungunya virus infection induces ADP-ribosylation through PARP1 activation, driven by caspase-3-mediated apoptosis, without transcriptionally activating PARPs. In contrast, astrocytic infections showed minimal ADP-ribosylation despite transcriptional activation of interferon-stimulated PARPs. Neurons exhibited limited innate immune response gene transcriptional activity, whereas astrocytes demonstrated strong upregulation of genes essential for pattern recognition receptor activation, thus enhancing double-stranded RNA sensing and increasing type I interferon production during infection. We posit that PARP1 activation and type I IFN response differentially regulate ADP-ribosylation in chikungunya virus-infected neural cells in a cell type-dependent manner. IMPORTANCE: Chikungunya virus is an emergent mosquito-borne alphavirus increasingly associated with neurological infection and subsequent long-term disabilities. Its continued global spread and recurrent outbreaks underscore its significant pandemic potential and the urgent need for effective countermeasures. Chikungunya virus showcases distinct, cell-type dependent replication dynamics within astrocytes and neurons, two major permissive cerebral cell types. However, understanding of the immunological basis of such cell type-specific infection dynamics remains limited, yet is necessary to elucidate virus pathogenesis within the brain and thus identification of downstream drug targets. Our study characterized two distinctly activated innate immunological pathways in chikungunya virus-infected astrocytes versus neurons, thus significantly contributing to molecular understanding cell type-specific chikungunya virus neurovirulence on a molecular level.