This study identifies critical sites like Madagascar, Somalia, Yemen, Mozambique, Sri Lanka, Myanmar, and the Maldives along five major bird migration routes (EAAF, AEF, CAF, MBSF, and EAAF)³, which facilitate the spread of AIV across continents⁵. Moreover, the consistent presence of H5N1 in ecologically critical sites underscores its ongoing circulation and potential adaptation in wild bird populations across tropical and subtropical zones. This pattern reinforces global concerns about H5N1’s expanding ecological footprint and the threats it poses to biodiversity and public health, especially along migratory bird pathways. The higher subtype richness per sample in Papua New Guinea, Indonesia, and the Philippines suggests a more heterogeneous viral population, potentially driven by diverse ecological niches, host species, or transmission interfaces. This diversity could reflect increased opportunities for viral reassortment or emergence of novel strains. Conversely, lower subtype richness in Yemen and Somalia may indicate more stable or homogenous transmission patterns. These regions serve as convergence zones for migratory birds from different geographical origins, potentially increasing the probability of exchange and reassortment of viral strains³. This highlights the importance of considering both ecological dynamics and avian movement patterns when interpreting patterns of viral diversity across sites. Several high-risk wild bird species contribute to avian influenza transmission across Myanmar, Papua New Guinea, Indonesia, the Philippines, Mozambique, Yemen, Somalia, Madagascar, Sri Lanka, and the Maldives. Waterfowl, including the Lesser Whistling Duck (Dendrocygna javanica) (Myanmar, Indonesia, Philippines, Sri Lanka, Madagascar), Northern Pintail (Anas acuta) (found in all sampled countries), Garganey (Spatula querquedula) (migratory across all these regions), and Pacific Black Duck (Anas superciliosa) (Papua New Guinea, Indonesia, Philippines), are known carriers of H5N1 and H9N2 and frequently mix with domestic ducks in flooded rice fields and wetlands, especially in rural farming communities^5,6. Shorebirds and waders, such as the Common Sandpiper (Actitis hypoleucos)(found in all sampled countries), Great Knot (Calidris tenuirostris) (Indonesia, Philippines, Myanmar), and Whimbrel (Numenius phaeopus) (long-distance migrant across all these regions), follow major flyways like the East Asian-Australasian, Central Asian, and East African Flyways, stopping at key wetlands, rice paddies, and coastal lagoons in these countries, where they may transmit AIV to local poultry and wild bird populations^5,6. Gulls and terns, including the Black-headed Gull (Chroicocephalus ridibundus) (Myanmar, Sri Lanka, Madagascar, Yemen) and Whiskered Tern (Chlidonias hybrida) (Indonesia, Philippines, Myanmar, Sri Lanka, Madagascar, Yemen), frequently scavenge at fish markets and coastal areas in low-income fishing communities, increasing disease spillover risks between wild birds, poultry, and humans⁶. Herons, egrets, and ibises, such as the Little Egret (Egretta garzetta)(all listed countries) and Glossy Ibis (Plegadis falcinellus) (Mozambique, Madagascar, Myanmar, Sri Lanka), inhabit rice paddies, irrigation canals, and marshlands, further intertwining wild and domestic bird populations^5,6,7. Birds of prey, including the Brahminy Kite (Haliastur indus) (Indonesia, Philippines, Papua New Guinea, Myanmar, Sri Lanka, Madagascar) and Eastern Marsh Harrier (Circus spilonotus) (Myanmar, Indonesia, Philippines, Madagascar), scavenge infected bird carcasses, sustaining AIV in both natural and human-modified landscapes⁶. Meanwhile, urban-adapted species like the Common Myna (Acridotheres tristis) (Myanmar, Sri Lanka, Maldives, Madagascar, Mozambique), Eurasian Tree Sparrow (Passer montanus) (Myanmar, Indonesia, Philippines, Sri Lanka, Mozambique), and House Crow (Corvus splendens) (Sri Lanka, Maldives, Myanmar, Mozambique, Yemen, Madagascar) spread the virus in small village communitas, informal markets, and backyard poultry farms, particularly where poultry waste, open food sources, and live bird trade create ideal conditions for viral persistence and mutation^5,6,7.

Fluctuations in AIV RNA concentrations appear to be shaped by a complex interplay of seasonal migration patterns, environmental conditions, and avian community composition. The temporal dynamics revealed a biphasic pattern, with two distinct periods of heightened viral activity: one centered around April–June and the other during December–February. These peaks likely correspond to seasonal ecological triggers, such as rainfall, food availability, and the congregation of migratory birds which are especially relevant in tropical and subtropical ecosystems⁵. This finding reinforces the importance of adopting regionally tailored seasonal frameworks in surveillance efforts, rather than applying temperate-biased assumptions.

Notably, the observed increases in AIV concentrations in countries like Madagascar, Yemen, and the Philippines between December–February 2021–2022 and April–June 2022, followed by widespread declines in August–October 2022, point toward episodic amplification events rather than continuous circulation. These brief but intense surges in viral burden suggest that transmission is likely driven by ecological pulses such as migratory staging or breeding events rather than persistent endemicity. The December–February 2022–2023 period marked another significant resurgence, with eight of ten countries recording their highest concentrations during this time, further supporting the role of seasonal ecological dynamics in viral transmission.

Spatial heterogeneity was also pronounced, with countries showing divergent patterns of increase and decline across seasons. This reflects localized risk factors such as habitat type, species diversity, climate, and proximity to migratory flyways^2,5. For example, countries positioned along key migratory corridors or at ecological convergence zones such as Yemen, Madagascar, and Sri Lanka tended to show higher variability and sharper seasonal peaks. Interestingly, AIV concentrations were lowest during August–October 2022, despite its alignment with major migratory movements in the EAAF, AEF, CAF, and MBSF. This unexpected dip may reflect a temporal lag between bird arrival and viral shedding, low bird densities, or suboptimal environmental conditions⁵ (e.g., temperature, humidity, and UV exposure), for virus persistence and detection during this period. Moreover, Many sampling sites were coastal or island habitats, where seawater exposure may inactivate AIV more rapidly than in freshwater environments⁵. Collectively, these findings underscore the dynamic, non-linear nature of AIV transmission in tropical and subtropical regions and highlight the need for flexible, frequent, and ecologically informed surveillance strategies.

Interestingly, we show that AIV RNA concentrations were highest during December–February 2022–2023, surpassing levels recorded in the same period of 2021–2022. This peak occurred before the widespread surge in avian influenza cases reported globally since 2023 until the percent^7,8,9. The subsequent global spread of the virus led to infections across multiple species, including wild and domestic birds, mammals, and even cases of spillover to humans^7,8,9. Moreover, although the Philippines officially reported its first H5N2 outbreak in backyard ducks in January 2025⁴, our data indicate that the virus may have been present in the region much earlier. This highlights potential gaps in existing surveillance efforts, where early circulation of AIV can go undetected. The observed pattern highlights the importance of continuous environmental surveillance in underrepresented regions to improve the detection of AIV circulation and mitigates the impact of avian influenza on both wildlife and public health⁷. The observed diversity in cleavage site sequences across different HA subtypes reflects the evolutionary strategies of AIV in adapting to various hosts and environmental conditions¹⁰. The presence of polybasic motifs in several H5 and H7 sequences raises concerns regarding the potential emergence of highly pathogenic strains, particularly in regions where domestic poultry and migratory birds coexist^7,10. The detection of these subtypes along multiple migratory flyways highlights the need for enhanced surveillance and genetic monitoring in this region.

Additionally, the detection of the H275Y mutation in H5N1-positive samples from the Bajuni Islands (Somalia), Socotra Archipelago (Yemen), and Maakandoodhoo (Maldives) raises critical concerns about the potential emergence of antiviral resistance in avian influenza viruses circulating in the wild. This mutation in the neuraminidase (NA) gene is associated with reduced sensitivity to oseltamivir, one of the few antiviral treatments available for severe influenza cases^11,12,13. Notably, these locations lie along major migratory bird flyways (CAF, MBSF, AEF and EAAF) and serve as key stopover points where birds from different regions intermingle. The convergence of high H5N1 prevalence, seasonal viral amplification, and resistance-associated mutations in these ecologically strategic sites underscores the pressing need for sustained surveillance. Strengthening monitoring efforts in such settings is essential not only for local public and animal health but also for anticipating the transboundary spread of drug-resistant AIV strains.

Additionally, the expansion of mining, logging, and infrastructure projects in Indonesia, Papua New Guinea, Madagascar, and Mozambique is pushing human settlements into remote forested regions and coastal islands, areas previously dominated by wild birds and migratory waterfowl. The increasing demand for nickel, cobalt, and rare earth metals to support the global electric vehicle (EV) revolution has led to rapid deforestation, wetland destruction, and the creation of artificial reservoirs and open-pit mines, which attract waterfowl and scavenging birds¹⁴. As workers, migrants, and displaced rural communities settle near these industrial sites, human-wildlife contact intensifies, increasing the risk of avian influenza spillover into domestic poultry and human populations^7,14.

At the same time, conflict zones in Myanmar, Somalia, and Yemen exacerbate the pandemic potential of HPAI. Ongoing civil unrest, displacement, and food insecurity have led to unregulated poultry trade, overcrowded refugee settlements, and weakened veterinary and public health infrastructure, all of which accelerate the risk of AIV transmission between humans, poultry, and wild birds¹⁵. In war-torn areas, military blockades, breakdowns in sanitation, and reliance on informal food sources force many communities to hunt wild birds or consume poorly regulated poultry, creating conditions for viral reassortment and the emergence of new AIV strains. Additionally, migratory birds that stop in these conflict zones, such as the Northern Pintail, Whimbrel, and Black-headed Gull, may act as silent carriers⁵, facilitating the long-distance spread of AI across unstable and poorly monitored regions.

This study provides some of the earliest available AIV surveillance data from countries like Sri Lanka, Myanmar, Papua New Guinea, Mozambique, Madagascar, Yemen, Somalia, and the Maldives. The presence of HPAI subtypes^1,9,16,17 like H5N1 in these regions highlights the need for targeted surveillance in migratory zones to mitigate spillover risks. A study conducted between 2006 and 2019, involving serological surveys of unvaccinated domestic ducks in Myanmar, revealed persistent yet intermittent circulation of H5 avian influenza viruses, even in years and regions without reported outbreaks¹⁸. This suggests silent or subclinical virus circulation in Myanmar and aligns with our findings. The diversity of detected clades, especially for H3N6 and H5N1, underscores the importance of global surveillance in tracking the spread and evolution of dominant and emerging subtypes². One of the key aims of this study was to highlight the glaring inequities in global infectious disease surveillance. Current systems are disproportionately concentrated in developed regions or limited to select locations or large-scale poultry farms, leaving vast areas of high zoonotic potential under-monitored. Our study spans a large geographical area with civil conflicts, ethnic strife, and political instability, providing a comprehensive overview of influenza virus diversity and spread in regions historically excluded from global surveillance networks. This study emphasizes the importance of these regions and demonstrates their role in shaping global AIV epidemiological patterns. While we acknowledge the limitations of only obtaining HA and NA gene sequences, the logistical and technical challenges of fieldwork in these remote settings often necessitate prioritization of the most epidemiologically informative genes. The HA and NA segments remain critical for monitoring vaccine strain efficacy and antigenic drift, which ensures the data retains its practical value despite the absence of complete genomes.

While this study highlights the potential role of tropical and subtropical regions in AIV detection, we caution against interpreting these areas as singular sources of viral emergence. AIV circulation is shaped by complex, multidirectional migratory and ecological dynamics, and no region should be framed as solely responsible for downstream outbreaks. Our goal is not to attribute origin or blame but rather to emphasize the value of equitable and regionally tailored surveillance systems, particularly in regions that have been historically under-monitored despite lying along key avian flyways. Strengthening global surveillance capacity is a shared priority that benefits all regions.

In conclusion, by incorporating targeted environmental surveillance in these high-risk locations, as demonstrated in this study, the detection of novel AIV subtypes can be achieved, ensuring timely global awareness and preparedness for emerging threats. The high inter-seasonal variability in several countries suggests that AIV risk is neither static nor uniform and underscores the value of integrating ecological, climatological, and behavioral data into viral surveillance frameworks. By aligning molecular findings with migratory ecology, we can move toward more predictive and targeted surveillance strategies that are better suited to dynamic, tropical systems.