Persistence of dengue serotype 2 viral RNA in blood cells of a returned traveler with dengue fever.

Dengue virus (DENV) is one of the most significant vector-borne pathogens worldwide. In this report, we describe clinical features and laboratory detection of dengue in a 45-year-old traveler to Nicaragua on return home to the United States in 2019. Clinical presentation was mild, with rash, headache, and fatigue, with only low-grade transient fever. Infection dynamics were documented by serology and PCR of serially collected body fluids. DENV serotype 2 was detected in whole blood 1 day after symptoms emerged, with viral RNA isolated to the red cell fraction, and remained detectable through day 89. DENV-2 RNA was detected in serum only on day 4, and IgM was undetectable on day 4 but evident by day 13. Viral RNA was also detected in urine. This report of DENV-2 RNA persistence in blood cells but only transient appearance in serum, supports the potential diagnostic value of whole blood over serum for PCR and opportunity of an expanded testing window. Informed testing approaches can improve diagnostic accuracy and inform strategies that preserve individual and public health.

Novel species of Triatoma (Hemiptera: Reduviidae) identified in a case of vectorial transmission of Chagas disease in northern Belize.

Chagas disease is a leading cause of non-ischemic cardiomyopathy in endemic regions of Central and South America. In Belize, Triatoma dimidiata sensu lato has been identified as the predominate taxon but vectorial transmission of Chagas disease is considered to be rare in the country. We recently identified an acute case of vector-borne Chagas disease in the northern region of Belize. Here we present a subsequent investigation of triatomines collected around the case-patient's home. We identified yet undescribed species, closely related to Triatoma huehuetenanguensis vector by molecular systematics methods occurring in the peridomestic environment. The identification of a T. cruzi-positive, novel species of Triatoma in Belize indicates an increased risk of transmission to humans in the region and warrants expanded surveillance and further investigation.

Impact of the COVID-19 pandemic on pediatric faculty: a report from nine academic institutions.

BACKGROUND: The COVID-19 pandemic affected home and work routines, which may exacerbate existing academic professional disparities. Objectives were to describe the impact of the pandemic on pediatric faculty's work productivity, identify groups at risk for widening inequities, and explore mitigation strategies. METHODS: A cross-sectional study of faculty members was conducted at nine U.S. pediatric departments. Responses were analyzed by demographics, academic rank, and change in home caregiving responsibility. RESULTS: Of 5791 pediatric faculty members eligible, 1504 (26%) completed the survey. The majority were female (64%), over 40 years old (60%), and assistant professors (47%). Only 7% faculty identified as underrepresented in medicine. Overall 41% reported an increase in caregiving during the pandemic. When comparing clinical, administrative, research, and teaching activities, faculty reported worse 1-year outlook for research activities. Faculty with increased caregiving responsibilities were more likely to report concerns over delayed promotion and less likely to have a favorable outlook regarding clinical and research efforts. Participants identified preferred strategies to mitigate challenges. CONCLUSIONS: The COVID-19 pandemic negatively impacted pediatric faculty productivity with the greatest effects on those with increased caregiving responsibilities. COVID-19 was particularly disruptive to research outlook. Mitigation strategies are needed to minimize the long-term impacts on academic pediatric careers. IMPACT: The COVID-19 pandemic most negatively impacted work productivity of academic pediatric faculty with caregiving responsibilities. COVID-19 was particularly disruptive to short-term (1-year) research outlook among pediatric faculty. Faculty identified mitigation strategies to minimize the long-term impacts of the pandemic on academic pediatric career pathways.

Proteome Analysis for Inflammation Related to Acute and Convalescent Infection.

Infectious diseases are a significant burden in global healthcare. Pathogens engage with different host defense mechanisms. However, it is currently unknown if there are disease-specific immune signatures and/or if different pathogens elicit common immune-associated molecular entities to common therapeutic interventions. We studied patients enrolled through the Human Immunology Project Consortium (HIPC), which focuses on immune responses to various infections. Blood samples were collected and analyzed from patients during infection and follow-up time points at the convalescent stage. The study included samples from patients with Lyme disease (LD), tuberculosis (TB), malaria (MLA), dengue virus (DENV), and West Nile virus (WNV), as well as kidney transplant patients with cytomegalovirus (CMV) and polyomavirus (BKV) infections. Using an antibody-based assay, we quantified ~ 350 cell surface markers, cytokines, and chemokines involved in inflammation and immunity. Unique protein signatures were identified specific to the acute phase of infection irrespective of the pathogen type, with significant changes during convalescence. In addition, tumor necrosis factor receptor superfamily member 6 (TNR6), C-C Motif Chemokine Receptor 7 (CCR7), and C-C motif chemokine ligand-1 (CCL1) were increased in the acute and convalescent phases across all viral, bacterial, and protozoan compared to blood from healthy donors. Furthermore, despite the differences between pathogens, proteins were enriched in common biological pathways such as cell surface receptor signaling pathway and response to external stimulus. In conclusion, we demonstrated that irrespective of the pathogen type, there are common immunoregulatory and proinflammatory signals.

Early cellular and molecular signatures correlate with severity of West Nile virus infection.

Infection with West Nile virus (WNV) drives a wide range of responses, from asymptomatic to flu-like symptoms/fever or severe cases of encephalitis and death. To identify cellular and molecular signatures distinguishing WNV severity, we employed systems profiling of peripheral blood from asymptomatic and severely ill individuals infected with WNV. We interrogated immune responses longitudinally from acute infection through convalescence employing single-cell protein and transcriptional profiling complemented with matched serum proteomics and metabolomics as well as multi-omics analysis. At the acute time point, we detected both elevation of pro-inflammatory markers in innate immune cell types and reduction of regulatory T cell activity in participants with severe infection, whereas asymptomatic donors had higher expression of genes associated with anti-inflammatory CD16+ monocytes. Therefore, we demonstrated the potential of systems immunology using multiple cell-type and cell-state-specific analyses to identify correlates of infection severity and host cellular activity contributing to an effective anti-viral response.

Blockade of interferon signaling decreases gut barrier integrity and promotes severe West Nile virus disease.

The determinants of severe disease caused by West Nile virus (WNV) and why only ~1% of individuals progress to encephalitis remain poorly understood. Here, we use human and mouse enteroids, and a mouse model of pathogenesis, to explore the capacity of WNV to directly infect gastrointestinal (GI) tract cells and contribute to disease severity. At baseline, WNV poorly infects human and mouse enteroid cultures and enterocytes in mice. However, when STAT1 or type I interferon (IFN) responses are absent, GI tract cells become infected, and this is associated with augmented GI tract and blood-brain barrier (BBB) permeability, accumulation of gut-derived molecules in the brain, and more severe WNV disease. The increased gut permeability requires TNF-α signaling, and is absent in WNV-infected IFN-deficient germ-free mice. To link these findings to human disease, we measured auto-antibodies against type I IFNs in serum from WNV-infected human cohorts. A greater frequency of auto- and neutralizing antibodies against IFN-α2 or IFN-ω is present in patients with severe WNV infection, whereas virtually no asymptomatic WNV-infected subjects have such antibodies (odds ratio 24 [95% confidence interval: 3.0 - 192.5; P = 0.003]). Overall, our experiments establish that blockade of type I IFN signaling extends WNV tropism to enterocytes, which correlates with increased gut and BBB permeability, and more severe disease.

Infectious profiles in pediatric anti-N-methyl-d-aspartate receptor encephalitis.

Anti-N-methyl-d-aspartate receptor autoimmune encephalitis (NMDAR AE) is an antibody-mediated neurological disorder that may be caused by post-herpes simplex virus-1 meningoencephalitis (HSV ME) and ovarian teratomas, although most pediatric cases are idiopathic. We sought to evaluate if other infections precede NMDAR AE by conducting a single-center, retrospective, case-control study of 86 pediatric cases presenting to Texas Children's Hospital between 2006 and 2022. HSV ME (HSV-1 and HSV-2) was a significantly more common preceding infection in the experimental group compared to control patients with idiopathic intracranial hypertension, while there was no difference in remote HSV infection between the two groups. Recent Epstein-Barr virus infection was evident in 8/42 (19%) tested experimental patients in comparison to 1/25 (4%) tested control patients which provided evidence for a genuine measure of effect but was not statistically significant due to small sample size (p = 0.07). The other 25 infectious etiologies were not different among the two groups and not all variables were clinically indicated or obtained in every subject, highlighting the need for future standardized, multi-institutional studies on underlying infectious precursors of autoimmune encephalitis.

Interleukins, Chemokines, and Tumor Necrosis Factor Superfamily Ligands in the Pathogenesis of West Nile Virus Infection.

West Nile virus (WNV) is a mosquito-borne pathogen that can lead to encephalitis and death in susceptible hosts. Cytokines play a critical role in inflammation and immunity in response to WNV infection. Murine models provide evidence that some cytokines offer protection against acute WNV infection and assist with viral clearance, while others play a multifaceted role WNV neuropathogenesis and immune-mediated tissue damage. This article aims to provide an up-to-date review of cytokine expression patterns in human and experimental animal models of WNV infections. Here, we outline the interleukins, chemokines, and tumor necrosis factor superfamily ligands associated with WNV infection and pathogenesis and describe the complex roles they play in mediating both protection and pathology of the central nervous system during or after virus clearance. By understanding of the role of these cytokines during WNV neuroinvasive infection, we can develop treatment options aimed at modulating these immune molecules in order to reduce neuroinflammation and improve patient outcomes.

Monitoring Temporal Changes in SARS-CoV-2 Spike Antibody Levels and Variant-Specific Risk for Infection, Dominican Republic, March 2021-August 2022.

To assess changes in SARS-CoV-2 spike binding antibody prevalence in the Dominican Republic and implications for immunologic protection against variants of concern, we prospectively enrolled 2,300 patients with undifferentiated febrile illnesses in a study during March 2021-August 2022. We tested serum samples for spike antibodies and tested nasopharyngeal samples for acute SARS-CoV-2 infection using a reverse transcription PCR nucleic acid amplification test. Geometric mean spike antibody titers increased from 6.6 (95% CI 5.1-8.7) binding antibody units (BAU)/mL during March-June 2021 to 1,332 (95% CI 1,055-1,682) BAU/mL during May-August 2022. Multivariable binomial odds ratios for acute infection were 0.55 (95% CI 0.40-0.74), 0.38 (95% CI 0.27-0.55), and 0.27 (95% CI 0.18-0.40) for the second, third, and fourth versus the first anti-spike quartile; findings were similar by viral strain. Combining serologic and virologic screening might enable monitoring of discrete population immunologic markers and their implications for emergent variant transmission.

Prevalence of Trypanosoma cruzi, the Etiologic Agent of Chagas Disease, Infection in Texas Skunks (Mammalia: Mephitidae).

Background: Chagas disease is one of the world's most neglected tropical diseases, infecting over six million people across the Americas. The hemoparasite Trypanosoma cruzi is the etiological agent for the disease, circulating in domestic, peridomestic, and sylvatic transmission cycles that are maintained by triatomine vectors and a diversity of wild and synanthropic hosts. Public health and wildlife management interventions targeting the interruption of T. cruzi transmission rely on an understanding of the dynamics driving the ecology of this zoonotic pathogen. One wildlife host that purportedly plays a role in the transmission of Chagas disease within the southern United States is the striped skunk (Mephitis mephitis), although infection prevalence in this species is poorly understood. Materials and Methods: To this end, we conducted a PCR-based surveillance of T. cruzi in 235 wild skunks, representing 4 species, across 76 counties and 10 ecoregions in Texas, United States, along with an evaluation of risk factors associated with the infection. Results: We recovered an overall T. cruzi prevalence of 17.9% for all mephitid taxa aggregated, ranging between 6.7% for plains spotted skunks (Spilogale putorius interrupta) and 42.9% for western spotted skunks (Spilogale gracilis). We report the first cases of T. cruzi infection in plains spotted and American hog-nosed skunks (Conepatus leuconotus), of important note for conservation medicine since populations of both species are declining within Texas. Although not statistically significant, we also detected trends for juveniles to exhibit greater infection risk than adults and for differential sex biases in T. cruzi prevalence between taxa, which align with variations in species-specific seasonal activity patterns. No geographic or taxonomic risk factors were identified. Conclusion: Our study contributed key data for population viability analyses and epidemiologic models in addition to providing a baseline for future T. cruzi surveillance among skunks and other wildlife species.

Protocol of Detection of West Nile Virus in Clinical Samples.

West Nile virus (WNV) is one of the leading causes of arboviral encephalitis in the United States but is often underdiagnosed. Despite the wide breadth of WNV-induced clinical disease syndromes, many of the symptoms associated with WNV are nonspecific at the time of presentation; thus, choosing the right diagnostic tool is essential to not only understand the true burden of disease but also provide pathogen-directed interventions for WNV-infected patients. In this chapter, we briefly discuss the three most common types of diagnostic methods for WNV in human clinical samples: nucleic acid detection, enzyme-linked immunoassay (ELISA), and plaque reduction neutralization test (PRNT) and present the method for PRNT.

Longitudinal serum proteomics analyses identify unique and overlapping host response pathways in Lyme disease and West Nile virus infection.

Advancement in proteomics methods for interrogating biological samples has helped identify disease biomarkers for early diagnostics and unravel underlying molecular mechanisms of disease. Herein, we examined the serum proteomes of 23 study participants presenting with one of two common arthropod-borne infections: Lyme disease (LD), an extracellular bacterial infection or West Nile virus infection (WNV), an intracellular viral infection. The LC/MS based serum proteomes of samples collected at the time of diagnosis and during convalescence were assessed using a depletion-based high-throughput shotgun proteomics (dHSP) pipeline as well as a non-depleting blotting-based low-throughput platform (MStern). The LC/MS integrated analyses identified host proteome responses in the acute and recovery phases shared by LD and WNV infections, as well as differentially abundant proteins that were unique to each infection. Notably, we also detected proteins that distinguished localized from disseminated LD and asymptomatic from symptomatic WNV infection. The proteins detected in both diseases with the dHSP pipeline identified unique and overlapping proteins detected with the non-depleting MStern platform, supporting the utility of both detection methods. Machine learning confirmed the use of the serum proteome to distinguish the infection from healthy control sera but could not develop discriminatory models between LD and WNV at current sample numbers. Our study is the first to compare the serum proteomes in two arthropod-borne infections and highlights the similarities in host responses even though the pathogens and the vectors themselves are different.

Incorporating COVID-19 into Acute Febrile Illness Surveillance Systems, Belize, Kenya, Ethiopia, Peru, and Liberia, 2020-2021.

Existing acute febrile illness (AFI) surveillance systems can be leveraged to identify and characterize emerging pathogens, such as SARS-CoV-2, which causes COVID-19. The US Centers for Disease Control and Prevention collaborated with ministries of health and implementing partners in Belize, Ethiopia, Kenya, Liberia, and Peru to adapt AFI surveillance systems to generate COVID-19 response information. Staff at sentinel sites collected epidemiologic data from persons meeting AFI criteria and specimens for SARS-CoV-2 testing. A total of 5,501 patients with AFI were enrolled during March 2020-October 2021; >69% underwent SARS-CoV-2 testing. Percentage positivity for SARS-CoV-2 ranged from 4% (87/2,151, Kenya) to 19% (22/115, Ethiopia). We show SARS-CoV-2 testing was successfully integrated into AFI surveillance in 5 low- to middle-income countries to detect COVID-19 within AFI care-seeking populations. AFI surveillance systems can be used to build capacity to detect and respond to both emerging and endemic infectious disease threats.

Diagnosis of Acute Chagas Disease in a Belizean Child with Evidence of a Multiclonal Trypanosoma cruzi Infection.

In January 2020, we instituted acute febrile illness surveillance in 11 hospitals and clinics across Belize. Within 3 months, we diagnosed an acute case of Chagas disease by polymerase chain reaction in a 7-year-old child in the northern part of the country. Phylogenetic analyses of the parasite from the acute blood specimen revealed a multiclonal Trypanosoma cruzi infection, including parasites from the TcII (25.0% of haplotypes), TcIV (2.5% of haplotypes), and TcV (72.5% of haplotypes) discrete typing units. The family reported no history of travel, and three Triatoma species vectors were found within the home. The child's mother was seronegative for antibodies to T. cruzi, ruling out congenital transmission. Convalescent blood samples documented seroconversion and confirmed acute infection. The child was successfully treated with nifurtimox. This is the first known diagnosed case of acute Chagas infection in Belize, highlighting the need for further investigation and public health prevention measures.

A Potential Role for Substance P in West Nile Virus Neuropathogenesis.

Of individuals who develop West Nile neuroinvasive disease (WNND), ~10% will die and >40% will develop long-term complications. Current treatment recommendations solely focus on supportive care; therefore, we urgently need to identify novel and effective therapeutic options. We observed a correlation between substance P (SP), a key player in neuroinflammation, and its receptor Neurokinin-1 (NK1R). Our study in a wild-type BL6 mouse model found that SP is upregulated in the brain during infection, which correlated with neuroinvasion and damage to the blood−brain barrier. Blocking the SP/NK1R interaction beginning at disease onset modestly improved survival and prolonged time to death in a small pilot study. Although SP is significantly increased in the brain of untreated WNND mice when compared to mock-infected animals, levels of WNV are unchanged, indicating that SP likely does not play a role in viral replication but may mediate the immune response to infection. Additional studies are necessary to define if SP plays a mechanistic role or if it represents other mechanistic pathways.

Potential Mechanisms Involved in Chronic Kidney Disease of Unclear Etiology.

BACKGROUND AND OBJECTIVES: The etiology of chronic kidney disease of unclear etiology, also known as Mesoamerican nephropathy, remains unclear. We investigated potential etiologies for Mesoamerican nephropathy in an immigrant dialysis population. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: Migrants with Mesoamerican nephropathy kidney failure (n=52) were identified by exclusion of known causes of kidney disease and compared using a cross-sectional survey with demographically similar patients with kidney failure from other causes (n=63) and age/sex/place of origin-matched healthy participants (n=16). Survey results were extended to the bench; C57BL/6 mice (n=73) received 10-15 weekly intraperitoneal injections of paraquat (a reactive oxygen species-generating herbicide) or vehicle. Kidney function, histology, and expression of organic cation transporter-2 (proximal tubule entry for paraquat) and multidrug and toxin extrusion 1 (extrusion pathway) were examined. Kidney biopsies from Nicaraguan patients with acute Mesoamerican nephropathy were stained for the above transporters and compared with patients with tubulointerstitial nephritis and without Mesoamerican nephropathy. RESULTS: Patients with Mesoamerican nephropathy and kidney failure were young agricultural workers, almost exclusively men; the majority were from Mexico and El Salvador; and they had prior exposures to agrochemicals, including paraquat (27%). After adjustment for age/sex, exposure to any agrochemical or paraquat was associated with Mesoamerican nephropathy kidney failure (odds ratio, 4.86; 95% confidence interval, 1.82 to 12.96; P=0.002 and odds ratio, 12.25; 95% confidence interval, 1.51 to 99.36; P=0.02, respectively). Adjusted for age/sex and other covariates, 1 year of agrochemical exposure was associated with Mesoamerican nephropathy kidney failure (odds ratio, 1.23; 95% confidence interval, 1.04 to 1.44; P=0.02). Compared with 16 matched healthy controls, Mesoamerican nephropathy kidney failure was significantly associated with exposure to paraquat and agrochemicals. Paraquat-treated male mice developed kidney failure and tubulointerstitial nephritis consistent with Mesoamerican nephropathy. Organic cation transporter-2 expression was higher in male kidneys versus female kidneys. Paraquat treatment increased organic cation transporter-2 expression and decreased multidrug and toxin extrusion 1 expression in male kidneys; similar results were observed in the kidneys of Nicaraguan patients with Mesoamerican nephropathy. CONCLUSIONS: Exposure to agrochemicals is associated with Mesoamerican nephropathy, and chronic exposure of mice to paraquat, a prototypical oxidant, induced kidney failure similar to Mesoamerican nephropathy.

A Preliminary Study of Proinflammatory Cytokines and Depression Following West Nile Virus Infection.

West Nile virus (WNV) is a neurotropic flavivirus that can cause acute febrile illness leading to neuroinvasive disease. Depression is a well-described outcome following infection, but the underlying pathogenic mechanisms are unknown. Proinflammatory cytokines play important roles in WNV infection, but their role in depression post-WNV remains unstudied. This research aimed to retrospectively evaluate associations between proinflammatory cytokines and new onset depression in a WNV cohort. Participants with asymptomatic WNV infection were significantly less likely to report new onset depression when compared to those with symptomatic disease. Participants with encephalitis and obesity were significantly more likely to report new onset depression post-infection. Based on univariate analysis of 15 antiviral or proinflammatory cytokines, depression was associated with elevated MCP-1 and decreased TNFα, whereas G-CSF was significantly elevated in those with a history of neuroinvasive WNV. However, no cytokines were statistically significant after adjusting for multiple comparisons using the Bonferroni method. While symptomatic WNV infection, encephalitis, and obesity were associated with new onset depression following infection, the role of proinflammatory cytokines requires additional studies. Further research involving paired acute-convalescent samples, larger sample sizes, and additional data points would provide additional insight into the impact of the inflammatory response on WNV-mediated depression.

Zoonotic Disease Testing Practices in Pediatric Patients with Meningitis and Encephalitis in a Subtropical Region.

Emerging vector-borne and zoonotic pathogens can cause neuroinvasive disease in children; utilization of appropriate diagnostic testing can be low, hindering diagnosis and clinical management of these cases. We must understand factors that influence healthcare providers' decisions to order diagnostic testing. We reviewed medical charts for pediatric meningitis and encephalitis patients (90 days-18 years) between 2010 and 2017 and analyzed variables associated with testing for known neuroinvasive zoonotic pathogens in the southern United States: West Nile virus (WNV), Bartonella spp., and Rickettsia spp. Among 620 cases of meningitis and encephalitis, ~1/3 (n = 209, 34%) were tested for WNV. Fewer cases were tested for Bartonella (n = 77, 12%) and Rickettsia (n = 47, 8%). Among those tested, 14 (7%) WNV, 7 (9%) Bartonella, and 6 (13%) Rickettsia cases were identified. Factors predicting testing were similar between all agents: clinical presentation of encephalitis, focal neurologic symptoms, new onset seizure, and decreased Glasgow Coma Scale on admission. Cases with a history of arthropod contact were more likely to be tested; however, we did not see an increase in testing during the summer season, when vector exposure typically increases. While our test utilization was higher than that reported in other studies, improvement is needed to identify zoonotic causes of neuroinvasive diseases.