The role of vaccination and environmental factors on outbreaks of high pathogenicity avian influenza H5N1 in Bangladesh.

High Pathogenicity Avian Influenza (HPAI) H5N1 outbreaks continue to wreak havoc on the global poultry industry and threaten the health of wild bird populations, with sporadic spillover in humans and other mammals, resulting in widespread calls to vaccinate poultry. Bangladesh has been vaccinating poultry since 2012, presenting a prime opportunity to study the effects of vaccination on HPAI H5N1circulation in both poultry and wild birds. We investigated the efficacy of vaccinating commercial poultry against HPAI H5N1 along with climatic and socio-economic factors considered potential drivers of HPAI H5N1 outbreak risk in Bangladesh. Using a multivariate modeling approach, we estimated that the rate of outbreaks was 18 times higher before compared to after vaccination, with winter months having a three times higher chance of outbreaks than summer months. Variables resulting in small but significant increases in outbreak rate were relatively low ambient temperatures for the time of year, literacy rate, chicken and duck density, crop density, and presence of highways; this may be attributable to low temperatures supporting viral survival outside the host, higher literacy driving reporting rate, density of the host reservoir, and spread of the virus through increased connectivity. Despite the substantial impact of vaccination on outbreaks, we note that HPAI H5N1 is still enzootic in Bangladesh; vaccinated poultry flocks have high rates of H5N1 prevalence, and spillover to wild birds has increased. Vaccination in Bangladesh thus bears the risk of supporting "silent spread," where the vaccine only provides protection against disease and not also infection. Our findings underscore that poultry vaccination can be part of holistic HPAI mitigation strategies when accompanied by monitoring to avoid silent spread.

In silico prediction of interaction between Nipah virus attachment glycoprotein and host cell receptors Ephrin-B2 and Ephrin-B3 in domestic and peridomestic mammals.

Nipah virus (NiV) is a lethal bat-borne zoonotic virus that causes mild to acute respiratory distress and neurological manifestations in humans with a high mortality rate. NiV transmission to humans occurs via consumption of bat-contaminated fruit and date palm sap (DPS), or through direct contact with infected individuals and livestock. Since NiV outbreaks were first reported in pigs from Malaysia and Singapore, non-neutralizing antibodies against NiV attachment Glycoprotein (G) have also been detected in a few domestic mammals. NiV infection is initiated after NiV G binds to the host cell receptors Ephrin-B2 and Ephrin-B3. In this study, we assessed the degree of NiV host tropism in domestic and peridomestic mammals commonly found in Bangladesh that may be crucial in the transmission of NiV by serving as intermediate hosts. We carried out a protein-protein docking analysis of NiV G complexes (n = 52) with Ephrin-B2 and B3 of 13 domestic and peridomestic species using bioinformatics tools. Protein models were generated by homology modelling and the structures were validated for model quality. The different protein-protein complexes in this study were stable, and their binding affinity (ΔG) scores ranged between -8.0 to -19.1 kcal/mol. NiV Bangladesh (NiV-B) strain displayed stronger binding to Ephrin receptors, especially with Ephrin-B3 than the NiV Malaysia (NiV-M) strain, correlating with the observed higher pathogenicity of NiV-B strains. From the docking result, we found that Ephrin receptors of domestic rat (R. norvegicus) had a higher binding affinity for NiV G, suggesting greater susceptibility to NiV infections compared to other study species. Investigations for NiV exposure to domestic/peridomestic animals will help us knowing more the possible role of rats and other animals as intermediate hosts of NiV and would improve future NiV outbreak control and prevention in humans and domestic animals.

Determinants for the presence of avian influenza virus in live bird markets in Bangladesh: Towards an easy fix of a looming one health issue.

Highly pathogenic avian influenza virus subtype H5N1 endangers poultry, wildlife, and human health and is enzootic in large parts of Asia, with live bird markets (LBMs) as putative hotspots for their maintenance, amplification, and spread. To mitigate the extent of these and other avian influenza viruses (AIV) of concern, we aimed to increase our quantitative understanding of the factors determining the presence of avian influenza virus in LBM stalls. Between 2016 and 2017, we collected fecal or offal samples from 1008 stalls in 113 LBMs across the Dhaka and Rajshahi districts in Bangladesh. For each stall, samples were pooled and tested for the AIV matrix gene, followed by H5 and H9 subtyping using rRT-PCR. We detected Influenza A viral RNA in 49% of the stalls. Of the AIV positive samples, 52% and 24% were determined to be H5 and H9 viruses, respectively, which are both subtypes of considerable health concern. We used generalized linear mixed effect modelling to study AIV presence in individual stalls within LBMs as a function of 13 out of the 20 risk factors identified by FAO. We found that small and feasible improvements in cleaning and disinfection frequency, installing running water in stalls, and not mixing different breeds of chicken in the same cages had large impacts on the presence of AIV in stalls (Odds ratios 0.03-0.05). Next, cleaning vehicles used in poultry transport, not selling waterfowl with chickens in the same stall, buying stock directly from commercial farms, separating sick birds from healthy ones, and avoiding access by wild birds like house crows, also had major effects on lowering the risk of stalls having AIV (Odds ratios 0.16-0.33). These findings can be directly used in developing practical and affordable measures to reduce the prevalence of AIV in LBMs. Also, in settings with limited resources like Bangladesh, such mitigation may significantly contribute to reducing AIV circulation amongst poultry and spillover to wildlife and humans.

Potential risk zones and climatic factors influencing the occurrence and persistence of avian influenza viruses in the environment of live bird markets in Bangladesh.

Live bird markets (LBMs) are critical for poultry trade in many developing countries that are regarded as hotspots for the prevalence and contamination of avian influenza viruses (AIV). Therefore, we conducted weekly longitudinal environmental surveillance in LBMs to determine annual cyclic patterns of AIV subtypes, environmental risk zones, and the role of climatic factors on the AIV presence and persistence in the environment of LBM in Bangladesh. From January 2018 to March 2020, we collected weekly fecal and offal swab samples from each LBM and tested using rRT-PCR for the M gene and subtyped for H5, H7, and H9. We used Generalized Estimating Equations (GEE) approaches to account for repeated observations over time to correlate the AIV prevalence and potential risk factors and the negative binomial and Poisson model to investigate the role of climatic factors on environmental contamination of AIV at the LBM. Over the study period, 37.8% of samples tested AIV positive, 18.8% for A/H5, and A/H9 was, for 15.4%. We found the circulation of H5, H9, and co-circulation of H5 and H9 in the environmental surfaces year-round. The Generalized Estimating Equations (GEE) model reveals a distinct seasonal pattern in transmitting AIV and H5. Specifically, certain summer months exhibited a substantial reduction of risk up to 70-90% and 93-94% for AIV and H5 contamination, respectively. The slaughtering zone showed a significantly higher risk of contamination with H5, with a three-fold increase in risk compared to bird-holding zones. From the negative binomial model, we found that climatic factors like temperature and relative humidity were also significantly associated with weekly AIV circulation. An increase in temperature and relative humidity decreases the risk of AIV circulation. Our study underscores the significance of longitudinal environmental surveillance for identifying potential risk zones to detect H5 and H9 virus co-circulation and seasonal transmission, as well as the imperative for immediate interventions to reduce AIV at LBMs in Bangladesh. We recommend adopting a One Health approach to integrated AIV surveillance across animal, human, and environmental interfaces in order to prevent the epidemic and pandemic of AIV.

Viral maintenance and excretion dynamics of coronaviruses within an Egyptian rousette fruit bat maternal colony: considerations for spillover.

Novel coronavirus species of public health and veterinary importance have emerged in the first two decades of the twenty-first century, with bats identified as natural hosts for progenitors of many coronaviruses. Targeted wildlife surveillance is needed to identify the factors involved in viral perpetuation within natural host populations, and drivers of interspecies transmission. We monitored a natural colony of Egyptian rousette bats at monthly intervals across two years to identify circulating coronaviruses, and to investigate shedding dynamics and viral maintenance within the colony. Three distinct lineages were detected, with different seasonal temporal excretion dynamics. For two lineages, the highest periods of coronavirus shedding were at the start of the year, when large numbers of bats were found in the colony. Highest peaks for a third lineage were observed towards the middle of the year. Among individual bat-level factors (age, sex, reproductive status, and forearm mass index), only reproductive status showed significant effects on excretion probability, with reproductive adults having lower rates of detection, though factors were highly interdependent. Analysis of recaptured bats suggests that viral clearance may occur within one month. These findings may be implemented in the development of risk reduction strategies for potential zoonotic coronavirus transmission.

Variations in small-scale movements of, Rousettus aegyptiacus, a Marburg virus reservoir across a seasonal gradient.

BACKGROUND: Bats are increasingly being recognized as important hosts for viruses, some of which are zoonotic and carry the potential for spillover within human and livestock populations. Biosurveillance studies focused on assessing the risk of pathogen transmission, however, have largely focused on the virological component and have not always considered the ecological implications of different species as viral hosts. The movements of known viral hosts are an important component for disease risk assessments as they can potentially identify regions of higher risk of contact and spillover. As such, this study aimed to synthesize data from both virological and ecological fields to provide a more holistic assessment of the risk of pathogen transmission from bats to people. RESULTS: Using radiotelemetry, we tracked the small-scale movements of Rousettus aegyptiacus, a species of bat known to host Marburg virus and other viruses with zoonotic potential, in a rural settlement in Limpopo Province, South Africa. The tracked bats exhibited seasonal variations in their movement patterns including variable usage of residential areas which could translate to contact between bats and humans and may facilitate spillover. We identified a trend for increased usage of residential areas during the winter months with July specifically experiencing the highest levels of bat activity within residential areas. July has previously been identified as a key period for increased spillover risk for viruses associated with R. aegyptiacus from this colony and paired with the increased activity levels, illustrates the risk for spillover to human populations. CONCLUSION: This study emphasizes the importance of incorporating ecological data such as movement patterns with virological data to provide a better understanding of the risk of pathogen spillover and transmission.

Prevalence and risk factors for avian influenza virus (H5 and H9) contamination in peri-urban and rural live bird markets in Bangladesh.

Avian influenza viruses (AIV) have been frequently detected in live bird markets (LBMs) around the world, primarily in urban areas, and have the ability to spillover to other species, including humans. Despite frequent detection of AIV in urban LBMs, the contamination of AIV on environmental surfaces in rural and peri-urban LBMs in Bangladesh is poorly documented. Therefore, we conducted this study to determine the prevalence of AIV subtypes within a subset of peri-urban and rural LBMs in Bangladesh and to further identify associated risk factors. Between 2017 and 2018, we collected faecal and offal samples from 200 stalls in 63 LBMs across four sub-districts. We tested the samples for the AIV matrix gene (M-gene) followed by H5, H7, and H9 subtypes using real-time reverse transcriptase-polymerase chain reaction (rRT-PCR). We performed a descriptive analysis of market cleanliness and sanitation practices in order to further elucidate the relationship between LBM biosecurity and AIV subtypes by species, sample types, and landscape. Subsequently, we conducted a univariate analysis and a generalized linear mixed model (GLMM) to determine the risk factors associated with AIV contamination at individual stalls within LBMs. Our findings indicate that practices related to hygiene and the circulation of AIV significantly differed between rural and peri-urban live bird markets. 42.5% (95% CI: 35.56-49.67) of stalls were positive for AIV. A/H5, A/H9, and A HA/Untyped were detected in 10.5% (95% CI: 6.62-15.60), 9% (95% CI: 5.42-13.85), and 24.0% (95% CI: 18.26-30.53) of stalls respectively, with no detection of A/H7. Significantly higher levels of AIV were found in the Sonali chicken strain compared to the exotic broiler, and in offal samples compared to fecal samples. In the GLMM analysis, we identified several significant risk factors associated with AIV contamination in LBMs at the stall level. These include: landscape (AOR: 3.02; 95% CI: 1.18-7.72), the number of chicken breeds present (AOR: 2.4; 95% CI: 1.01-5.67), source of birds (AOR: 2.35; 95% CI: 1.0-5.53), separation of sick birds (AOR: 3.04; 95% CI: 1.34-6.92), disposal of waste/dead birds (AOR: 3.16; 95% CI: 1.41-7.05), cleaning agent (AOR: 5.99; 95% CI: 2.26-15.82), access of dogs (AOR: 2.52; 95% CI: 1.12-5.7), wild birds observed on site (AOR: 2.31; 95% CI: 1.01-5.3). The study further revealed a substantial prevalence of AIV with H5 and H9 subtypes in peri-urban and rural LBMs. The inadequate biosecurity measures at poultry stalls in Bangladesh increase the risk of AIV transmission from poultry to humans. To prevent the spread of AIV to humans and wild birds, we suggest implementing regular surveillance at live bird markets and enhancing biosecurity practices in peri-urban and rural areas in Bangladesh.

Metagenomics-enabled reverse-genetics assembly and characterization of myotis bat morbillivirus.

Morbilliviruses are among the most contagious viral pathogens of mammals. Although previous metagenomic surveys have identified morbillivirus sequences in bats, full-length morbilliviruses from bats are limited. Here we characterize the myotis bat morbillivirus (MBaMV) from a bat surveillance programme in Brazil, whose full genome was recently published. We demonstrate that the fusion and receptor binding protein of MBaMV utilize bat CD150 and not human CD150, as an entry receptor in a mammalian cell line. Using reverse genetics, we produced a clone of MBaMV that infected Vero cells expressing bat CD150. Electron microscopy of MBaMV-infected cells revealed budding of pleomorphic virions, a characteristic morbillivirus feature. MBaMV replication reached 103-105 plaque-forming units ml-1 in human epithelial cell lines and was dependent on nectin-4. Infection of human macrophages also occurred, albeit 2-10-fold less efficiently than measles virus. Importantly, MBaMV is restricted by cross-neutralizing human sera elicited by measles, mumps and rubella vaccination and is inhibited by orally bioavailable polymerase inhibitors in vitro. MBaMV-encoded P/V genes did not antagonize human interferon induction. Finally, we show that MBaMV does not cause disease in Jamaican fruit bats. We conclude that, while zoonotic spillover into humans may theoretically be plausible, MBaMV replication would probably be controlled by the human immune system.

Characterization and epidemiology of antimicrobial resistance patterns of Salmonella spp. and Staphylococcus spp. in free-ranging rhesus macaque (Macaca mulatta) at high-risk interfaces with people and livestock in Bangladesh.

Introduction: Antimicrobial resistance (AMR) is a growing global health threat for humans and animals. Environmental contamination of antimicrobials from human and domestic animal feces has been linked to AMR in wildlife populations, including rhesus macaques. This study aimed to describe the eco-epidemiology of AMR within Salmonella and Staphylococcus species isolated from rhesus macaques. Methods: We followed macaque groups for 4 h per day (2 days) to observe the direct and indirect contact rate and type between macaques and people and livestock. We collected 399 freshly defecated, non-invasive fecal samples from macaques at seven sites in Bangladesh in January-June 2017. Bacterial isolation and identification were conducted using culture, biochemical characteristics, and polymerase chain reaction (PCR). An antimicrobial susceptibility test (AST) for 12 antimicrobials for each organism was conducted using the Kirby-Bauer disc diffusion method. Results: The overall prevalence of Salmonella spp. and Staphylococcus spp. in rhesus macaques was 5% (n = 18; 95% CI: 3-7%) and 16% (n = 64; 95% CI: 13-20%), respectively. All the isolated Salmonella spp. and most of the Staphylococcus spp. (95%; 61/64; 95% CI: 86.9-99%) were resistant to at least one antimicrobial. The odds of a fecal sample having antimicrobial-resistant Salmonella spp (OR = 6.6; CI: 0.9-45.8, P = 0.05) and Staphylococcus spp. (OR = 5.6; CI: 1.2-26, P = 0.02) were significantly higher in samples collected at peri-urban sites than those collected at rural and urban sites. Salmonella spp. were most frequently resistant to tetracycline (89%), azithromycin (83%), sulfamethoxazole-trimethoprim (50%), and nalidixic acid (44%). Staphylococcus spp. were found to be highly resistant to ampicillin (93%), methicillin (31%), clindamycin (26%), and rifampicin (18%). Both bacterial species produced colonies with multidrug resistance to up to seven antimicrobials. Direct and indirect contact rates (within 20 m for at least 15 min) and resource sharing between macaques and people were higher in urban sites, while macaque-livestock contact rates were higher in rural sites. Discussion: The study shows that resistant microorganisms are circulating in rhesus macaque, and direct and indirect contact with humans and livestock might expand the resistant organisms.

Interventions to Reduce Risk for Pathogen Spillover and Early Disease Spread to Prevent Outbreaks, Epidemics, and Pandemics.

The pathogens that cause most emerging infectious diseases in humans originate in animals, particularly wildlife, and then spill over into humans. The accelerating frequency with which humans and domestic animals encounter wildlife because of activities such as land-use change, animal husbandry, and markets and trade in live wildlife has created growing opportunities for pathogen spillover. The risk of pathogen spillover and early disease spread among domestic animals and humans, however, can be reduced by stopping the clearing and degradation of tropical and subtropical forests, improving health and economic security of communities living in emerging infectious disease hotspots, enhancing biosecurity in animal husbandry, shutting down or strictly regulating wildlife markets and trade, and expanding pathogen surveillance. We summarize expert opinions on how to implement these goals to prevent outbreaks, epidemics, and pandemics.

Nipah Virus Exposure in Domestic and Peridomestic Animals Living in Human Outbreak Sites, Bangladesh, 2013-2015.

Spillovers of Nipah virus (NiV) from Pteropus bats to humans occurs frequently in Bangladesh, but the risk for spillover into other animals is poorly understood. We detected NiV antibodies in cattle, dogs, and cats from 6 sites where spillover human NiV infection cases occurred during 2013-2015.

Lessons from the pandemic: Responding to emerging zoonotic viral diseases-a Keystone Symposia report.

The COVID-19 pandemic caught the world largely unprepared, including scientific and policy communities. On April 10-13, 2022, researchers across academia, industry, government, and nonprofit organizations met at the Keystone symposium "Lessons from the Pandemic: Responding to Emerging Zoonotic Viral Diseases" to discuss the successes and challenges of the COVID-19 pandemic and what lessons can be applied moving forward. Speakers focused on experiences not only from the COVID-19 pandemic but also from outbreaks of other pathogens, including the Ebola virus, Lassa virus, and Nipah virus. A general consensus was that investments made during the COVID-19 pandemic in infrastructure, collaborations, laboratory and manufacturing capacity, diagnostics, clinical trial networks, and regulatory enhancements-notably, in low-to-middle income countries-must be maintained and strengthened to enable quick, concerted responses to future threats, especially to zoonotic pathogens.

Predicting the potential for zoonotic transmission and host associations for novel viruses.

Host-virus associations have co-evolved under ecological and evolutionary selection pressures that shape cross-species transmission and spillover to humans. Observed virus-host associations provide relevant context for newly discovered wildlife viruses to assess knowledge gaps in host-range and estimate pathways for potential human infection. Using models to predict virus-host networks, we predicted the likelihood of humans as hosts for 513 newly discovered viruses detected by large-scale wildlife surveillance at high-risk animal-human interfaces in Africa, Asia, and Latin America. Predictions indicated that novel coronaviruses are likely to infect a greater number of host species than viruses from other families. Our models further characterize novel viruses through prioritization scores and directly inform surveillance targets to identify host ranges for newly discovered viruses.

Classification of new morbillivirus and jeilongvirus sequences from bats sampled in Brazil and Malaysia.

As part of a broad One Health surveillance effort to detect novel viruses in wildlife and people, we report several paramyxovirus sequences sampled primarily from bats during 2013 and 2014 in Brazil and Malaysia, including seven from which we recovered full-length genomes. Of these, six represent the first full-length paramyxovirid genomes sequenced from the Americas, including two that are the first full-length bat morbillivirus genome sequences published to date. Our findings add to the vast number of viral sequences in public repositories, which have been increasing considerably in recent years due to the rising accessibility of metagenomics. Taxonomic classification of these sequences in the absence of phenotypic data has been a significant challenge, particularly in the subfamily Orthoparamyxovirinae, where the rate of discovery of novel sequences has been substantial. Using pairwise amino acid sequence classification (PAASC), we propose that five of these sequences belong to members of the genus Jeilongvirus and two belong to members of the genus Morbillivirus. We also highlight inconsistencies in the classification of Tupaia virus and Mòjiang virus using the same demarcation criteria and suggest reclassification of these viruses into new genera. Importantly, this study underscores the critical importance of sequence length in PAASC analysis as well as the importance of biological characteristics such as genome organization in the taxonomic classification of viral sequences.

Nipah Virus Detection at Bat Roosts after Spillover Events, Bangladesh, 2012-2019.

Knowledge of the dynamics and genetic diversity of Nipah virus circulating in bats and at the human-animal interface is limited by current sampling efforts, which produce few detections of viral RNA. We report a series of investigations at Pteropus medius bat roosts identified near the locations of human Nipah cases in Bangladesh during 2012-2019. Pooled bat urine was collected from 23 roosts; 7 roosts (30%) had >1 sample in which Nipah RNA was detected from the first visit. In subsequent visits to these 7 roosts, RNA was detected in bat urine up to 52 days after the presumed exposure of the human case-patient, although the probability of detection declined rapidly with time. These results suggest that rapidly deployed investigations of Nipah virus shedding from bat roosts near human cases could increase the success of viral sequencing compared with background surveillance and could enhance understanding of Nipah virus ecology and evolution.

URSID γ-HERPESVIRUS TYPE 1-RELATED VIRUS IN CAPTIVE BORNEAN SUN BEARS (HELARCTOS MALAYANUS EURYSPILUS) IN SABAH, MALAYSIA.

The Bornean sun bear (Helarctos malayanus euryspilus) is the smallest subspecies of sun bear. Their numbers are declining, and more research is needed to better understand their health and biology. Forty-four bears housed at the Bornean Sun Bear Conservation Centre (BSBCC) in Sabah, Malaysia, were screened for known and novel viruses in November 2018. Ursid γ-herpesvirus type 1 (UrHV-1) is a herpesvirus that has been detected from swab samples of clinically healthy sun bears and biopsy samples of oral squamous cell carcinoma in sun bears. We detected an UrHV-1-related virus from throat and rectal swabs by molecular viral screening in samples from 15.9% of the sun bears at BSBCC. None of the bears with the UrHV-1-related virus in this study had oral lesions. There is no known report of UrHV-1 detection in the wild sun bear population, and its association with oral squamous cell carcinoma is not fully understood. Finding an UrHV-1-related virus in a rehabilitation center is a concern because conditions in captivity may contribute to spreading this virus, and there is the potential of introducing it into wild populations when a bear is released. This study demonstrates an urgent need to carry out similar surveillance for sun bears in captivity as well as those in the wild, to better understand the impact of captivity on the prevalence and spread of UrHV-1-related viruses. Positive bears also should be monitored for oral lesions to better understand whether there is a causal relationship.

Living Safely With Bats: Lessons in Developing and Sharing a Global One Health Educational Resource.

As part of a public health behavior change and communication strategy related to the identification of a novel ebolavirus in bats in Sierra Leone in 2016, a consortium of experts launched an effort to create a widely accessible resource for community awareness and education on reducing disease risk. The resulting picture book, Living Safely With Bats, includes technical content developed by a consortium of experts in public health, animal health, conservation, bats, and disease ecology from 30 countries. The book has now been adapted, translated, and used in more than 20 countries in Africa and Asia. We review the processes used to integrate feedback from local stakeholders and multidisciplinary experts. We also provide recommendations for One Health and other practitioners who choose to pursue the development and evaluation of this or similar zoonotic disease risk mitigation tools.

Seasonal shedding patterns of diverse henipavirus-related paramyxoviruses in Egyptian rousette bats.

Bat-borne viruses in the Henipavirus genus have been associated with zoonotic diseases of high morbidity and mortality in Asia and Australia. In Africa, the Egyptian rousette bat species (Rousettus aegyptiacus) is an important viral host in which Henipavirus-related viral sequences have previously been identified. We expanded these findings by assessing the viral dynamics in a southern African bat population. A longitudinal study of henipavirus diversity and excretion dynamics identified 18 putative viral species circulating in a local population, three with differing seasonal dynamics, and the winter and spring periods posing a higher risk of virus spillover and transmission. The annual peaks in virus excretion are most likely driven by subadults and may be linked to the waning of maternal immunity and recolonization of the roost in early spring. These results provide insightful information into the bat-host relationship that can be extrapolated to other populations across Africa and be communicated to at-risk communities as a part of evidence-based public health education and prevention measures against pathogen spillover threats.

Environmental Change and Zoonotic Disease Risk at Human-Macaque Interfaces in Bangladesh.

Anthropogenic land-use changes increase the frequency of interactions and habitat overlap between humans and macaques which play an important role in zoonotic disease transmission. This exploratory qualitative study aimed to examine connections between land-use change and macaque-human interactions and assess the chance of zoonotic disease transmission. We conducted ethnographic interviews and focus group discussions in Old Dhaka, Madaripur, and Chandpur, Bangladesh. Participants reported significant anthropogenic landscape transformations leading to increased human-macaque contact in the study areas. Participants also reported that all three sites underwent substantial landscape alteration from natural or agricultural land to a human-altered environment with roads, commercial, and residential buildings. Participants noted that the disappearance of forestland appeared to increase the macaque dependence on backyard fruit trees. Where rivers and ponds were filled to support local construction, macaques were also observed as becoming more dependent upon human water sources. These changed may help expanding the macaques' foraging areas, and they appear to be invading new areas where people are not culturally habituated to living with them. In response, many residents reported reacting aggressively toward the macaques, which they believed led to more bites and scratches. However, other respondents accepted the presence of macaques around their homes. Few participants considered macaques to be a source of disease transmission. This study revealed that local environmental changes, deforestation, urban expansion, construction, and water bodies' disappearance are linked to increasing human-macaque interactions. Understanding these interactions is critical to develop successful mitigation interventions at interfaces with a high risk for viral disease spillover.