Quantifying heterogeneities in arbovirus transmission: Description of the rationale and methodology for a prospective longitudinal study of dengue and Zika virus transmission in Iquitos, Peru (2014-2019).

Current knowledge of dengue virus (DENV) transmission provides only a partial understanding of a complex and dynamic system yielding a public health track record that has more failures than successes. An important part of the problem is that the foundation for contemporary interventions includes a series of longstanding, but untested, assumptions based on a relatively small portion of the human population; i.e., people who are convenient to study because they manifest clinically apparent disease. Approaching dengue from the perspective of people with overt illness has produced an extensive body of useful literature. It has not, however, fully embraced heterogeneities in virus transmission dynamics that are increasingly recognized as key information still missing in the struggle to control the most important insect-transmitted viral infection of humans. Only in the last 20 years have there been significant efforts to carry out comprehensive longitudinal dengue studies. This manuscript provides the rationale and comprehensive, integrated description of the methodology for a five-year longitudinal cohort study based in the tropical city of Iquitos, in the heart of the Peruvian Amazon. Primary data collection for this study was completed in 2019. Although some manuscripts have been published to date, our principal objective here is to support subsequent publications by describing in detail the structure, methodology, and significance of a specific research program. Our project was designed to study people across the entire continuum of disease, with the ultimate goal of quantifying heterogeneities in human variables that affect DENV transmission dynamics and prevention. Because our study design is applicable to other Aedes transmitted viruses, we used it to gain insights into Zika virus (ZIKV) transmission when during the project period ZIKV was introduced and circulated in Iquitos. Our prospective contact cluster investigation design was initiated by detecttion of a person with a symptomatic DENV infection and then followed that person's immediate contacts. This allowed us to monitor individuals at high risk of DENV infection, including people with clinically inapparent and mild infections that are otherwise difficult to detect. We aimed to fill knowledge gaps by defining the contribution to DENV transmission dynamics of (1) the understudied majority of DENV-infected people with inapparent and mild infections and (2) epidemiological, entomological, and socio-behavioral sources of heterogeneity. By accounting for factors underlying variation in each person's contribution to transmission we sought to better determine the type and extent of effort needed to better prevent virus transmission and disease.

The impact of dengue illness on social distancing and caregiving behavior.

BACKGROUND: Human mobility among residential locations can drive dengue virus (DENV) transmission dynamics. Recently, it was shown that individuals with symptomatic DENV infection exhibit significant changes in their mobility patterns, spending more time at home during illness. This change in mobility is predicted to increase the risk of acquiring infection for those living with or visiting the ill individual. It has yet to be considered, however, whether social contacts are also changing their mobility, either by socially distancing themselves from the infectious individual or increasing contact to help care for them. Social, or physical, distancing and caregiving could have diverse yet important impacts on DENV transmission dynamics; therefore, it is necessary to better understand the nature and frequency of these behaviors including their effect on mobility. METHODOLOGY AND PRINCIPAL FINDINGS: Through community-based febrile illness surveillance and RT-PCR infection confirmation, 67 DENV positive (DENV+) residents were identified in the city of Iquitos, Peru. Using retrospective interviews, data were collected on visitors and home-based care received during the illness. While 15% of participants lost visitors during their illness, 22% gained visitors; overall, 32% of all individuals (particularly females) received visitors while symptomatic. Caregiving was common (90%), particularly caring by housemates (91%) and caring for children (98%). Twenty-eight percent of caregivers changed their behavior enough to have their work (and, likely, mobility patterns) affected. This was significantly more likely when caring for individuals with low "health-related quality of well-being" during illness (Fisher's Exact, p = 0.01). CONCLUSIONS/SIGNIFICANCE: Our study demonstrates that social contacts of individuals with dengue modify their patterns of visitation and caregiving. The observed mobility changes could impact a susceptible individual's exposure to virus or a presymptomatic/clinically inapparent individual's contribution to onward transmission. Accounting for changes in social contact mobility is imperative in order to get a more accurate understanding of DENV transmission.

Heterogeneity of Dengue Illness in Community-Based Prospective Study, Iquitos, Peru.

Measuring heterogeneity of dengue illness is necessary to define suitable endpoints in dengue vaccine and therapeutic trials and will help clarify behavioral responses to illness. To quantify heterogeneity in dengue illness, including milder cases, we developed the Dengue Illness Perceptions Response (IPR) survey, which captured detailed symptom data, including intensity, duration, and character, and change in routine activities caused by illness. During 2016-2019, we collected IPR data daily during the acute phase of illness for 79 persons with a positive reverse transcription PCR result for dengue virus RNA. Most participants had mild ambulatory disease. However, we measured substantial heterogeneity in illness experience, symptom duration, and maximum reported intensity of individual symptoms. Symptom intensity was a more valuable predicter of major activity change during dengue illness than symptom presence or absence alone. These data suggest that the IPR measures clinically useful heterogeneity in dengue illness experience and its relation to altered human behavior.

Measuring health related quality of life for dengue patients in Iquitos, Peru.

Previous studies measuring the health-related quality of life (HRQoL) of individuals with dengue focused on treatment seeking populations. However, the vast majority of global dengue cases are unlikely to be detected by health systems. Representative measurements of HRQoL should therefore include patients with disease not likely to trigger treatment-seeking behavior. This study based in Iquitos, Peru used the Quality of Wellbeing Scale-Self Administered, a survey that enquires about not only physical health, but also psychological health, self-care, mobility, and usual social activities, and rates HRQoL between 0 (death) and 1 (optimum function), to evaluate the impact of dengue on HRQoL. In order to enroll treatment and non treatment-seeking participants, three modalities of participant recruitment were used. In addition to clinic and community-based febrile surveillance, a contact-cluster methodology was also employed to identify infected individuals less likely to seek treatment. We measured changes in HRQoL and identified common areas of health impairment in 73 virologically confirmed dengue cases at 3 time points during the participant's illness; the early-acute (days 0-6 post symptom onset), late-acute (days 7-20), and convalescent illness phases (days 21 +). Participants reported HRQoL related impairments at significantly higher frequency during the early-acute versus convalescent illness phase (Fisher's exact: P<0.01). There was substantial heterogeneity in scores during each illness phase with median scores in the early-acute, late-acute and convalescent phases of 0.56 (IQR: 0.41-0.64), 0.70 (IQR: 0.57-0.94), and 1 (IQR: 0.80-1.00), respectively. In all illness phases participants recruited in clinics had on average the lowest HRQoL scores where as those recruited in the contact clusters had the highest. Only 1 individual who was recruited in the contact-clusters had no reduction in HRQoL score during their illness. These data illustrate that dengue should be considered as a disease that may have significant implications for not only physical health but also psychological health and social functioning. The impact of dengue on the HRQoL of non-treatment-seeking individuals, although lower than the impact among treatment-seeking individuals, is not necessarily trivial.

Correction: Dengue illness impacts daily human mobility patterns in Iquitos, Peru.

[This corrects the article DOI: 10.1371/journal.pntd.0007756.].

Dengue illness impacts daily human mobility patterns in Iquitos, Peru.

BACKGROUND: Human mobility plays a central role in shaping pathogen transmission by generating spatial and/or individual variability in potential pathogen-transmitting contacts. Recent research has shown that symptomatic infection can influence human mobility and pathogen transmission dynamics. Better understanding the complex relationship between symptom severity, infectiousness, and human mobility requires quantification of movement patterns throughout infectiousness. For dengue virus (DENV), human infectiousness peaks 0-2 days after symptom onset, making it paramount to understand human movement patterns from the beginning of illness. METHODOLOGY AND PRINCIPAL FINDINGS: Through community-based febrile surveillance and RT-PCR assays, we identified a cohort of DENV+ residents of the city of Iquitos, Peru (n = 63). Using retrospective interviews, we measured the movements of these individuals when healthy and during each day of symptomatic illness. The most dramatic changes in mobility occurred during the first three days after symptom onset; individuals visited significantly fewer locations (Wilcoxon test, p = 0.017) and spent significantly more time at home (Wilcoxon test, p = 0.005), compared to when healthy. By 7-9 days after symptom onset, mobility measures had returned to healthy levels. Throughout an individual's symptomatic period, the day of illness and their subjective sense of well-being were the most significant predictors for the number of locations and houses they visited. CONCLUSIONS/SIGNIFICANCE: Our study is one of the first to collect and analyze human mobility data at a daily scale during symptomatic infection. Accounting for the observed changes in human mobility throughout illness will improve understanding of the impact of disease on DENV transmission dynamics and the interpretation of public health-based surveillance data.

Zika Virus: The Agent and Its Biology, With Relevance to Pathology.

Once obscure, Zika virus (ZIKV) has attracted significant medical and scientific attention in the past year because of large outbreaks associated with the recent introduction of this virus into the Western hemisphere. In particular, the occurrence of severe congenital infections and cases of Guillain-Barré syndrome has placed this virus squarely in the eyes of clinical and anatomic pathologists. This review article provides a basic introduction to ZIKV, its genetics, its structural characteristics, and its biology. A multidisciplinary effort will be essential to establish clinicopathologic correlations of the basic virology of ZIKV in order to advance development of diagnostics, therapeutics, and vaccines.

Gene expression profiling of dengue infected human primary cells identifies secreted mediators in vivo.

We used gene expression profiling of human primary cells infected in vitro with dengue virus (DENV) as a tool to identify secreted mediators induced in response to the infection. Affymetrix GeneChip analysis of human primary monocytes, B cells and dendritic cells infected with DENV in vitro showed strong induction of monocyte chemotactic protein 2 (MCP-2/CCL8), interferon gamma-induced protein 10 (IP-10/CXCL10) and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL/TNFSF10). The expression of these genes was confirmed in dendritic cells infected with DENV in vitro at mRNA and protein levels. A prospectively enrolled cohort of DENV-infected Venezuelan patients was used to measure the levels of these proteins in serum during three different periods of the disease. Results showed significant increase of MCP-2, IP-10, and TRAIL levels in patients infected with DENV during the febrile period, when compared to healthy donors and patients with other febrile illnesses. MCP-2 and IP-10 levels were still elevated during the post-febrile period while TRAIL levels dropped close to normal after defervescense. Patients with primary infections had higher TRAIL levels than patients with secondary infections during the febrile period of the disease. Increased levels of IP-10, TRAIL and MCP-2 in acute DENV infections suggest a role for these mediators in the immune response to the infection. MCP-2 was identified in this work as a new unreported and important dengue-related protein and IP-10 was confirmed as a novel and strong pro-inflammatory marker in acute disease.

Evidence of vascular damage in dengue disease: demonstration of high levels of soluble cell adhesion molecules and circulating endothelial cells.

Clinical evidence suggests that vascular damage plays a key role in the pathophysiology of dengue hemorrhagic fever (DHF). In this study, the authors tested this hypothesis by examining the levels of soluble intercellular adhesion molecule and vascular cell adhesion molecule (sICAM-1 and sVCAM-1), and the presence of circulating endothelial cells (CECs), as evidence of vascular damage, in peripheral blood from DHF patients (n=13). A significant increase in plasma levels of sICAM-1 (n=12) and sVCAM-1 (n=13) was detected by enzyme-linked immunosorbent assay (ELISA) in DHF patients, compared with healthy individuals. Increased numbers of CECs, as detected by the expression of endothelial cell markers (ICAM-1, platelet cell adhesion molecule [PCAM]-1, and CD36) with flow cytometry, were observed in DHF patients (n=4), compared to healthy subjects. The high levels of sICAM-1 and sVCAM-1, together with the presence of CECs in DHF patients, provide further evidence of endothelium damage and activation in DHF patients.

Relationship of thrombopoietin and interleukin-11 levels to thrombocytopenia associated with dengue disease.

Thrombocytopenia is one of the main clinical findings of dengue. In this work we examined the levels of thrombopoietin (TPO) and interleukin-11 (IL-11), two of the most potent regulators of platelet production, in serum from 28 patients with dengue fever (DF). Patients with DF had increased levels of TPO, compared with healthy individuals (p<0.005). Patients with dengue hemorrhagic fever (DHF, n=7), the more severe form of dengue, had higher TPO levels than patients with DF (p<0.001). Serum TPO levels and platelet counts were inversely correlated in both DF and DHF patients. IL-11 was detectable in neither DF nor DHF patients. Our results demonstrate that thrombocytopenia in dengue disease is associated with changes in the serum levels of TPO, but not IL-11, suggesting that this cytokine could be a potential early clinical marker of the severity of dengue disease.

Proinflammatory factors present in sera from patients with acute dengue infection induce activation and apoptosis of human microvascular endothelial cells: possible role of TNF-alpha in endothelial cell damage in dengue.

There is evidence that severe dengue disease is associated with alterations of the microvascular endothelium. We examined the hypothesis that activation and damage of microvascular endothelial cells (EC) could be induced by inflammatory mediators present in dengue patient's sera. We cultured human microvascular EC (HMEC-1) in vitro with sera from patients with acute dengue infection. Sera from patients with acute dengue induced an increase in ICAM-1 expression on HMEC-1. This effect was greater with samples from the acute febrile phase than with samples from the convalescent phase of the disease. Acute dengue sera had elevated levels of TNF-alpha and the endothelial activating effect of acute dengue sera was inhibited up to 80% by pre-treatment with monoclonal antibodies against TNF-alpha. Furthermore, acute dengue sera induced apoptosis in HMEC-1. These findings support the pathophysiologic significance of microvascular EC and serum inflammatory mediators in dengue.