Revisiting the Woolly wolf (Canis lupus chanco) phylogeny in Himalaya: Addressing taxonomy, spatial extent and distribution of an ancient lineage in Asia.

Of the sub-species of Holarctic wolf, the Woolly wolf (Canis lupus chanco) is uniquely adapted to atmospheric hypoxia and widely distributed across the Himalaya, Qinghai Tibetan Plateau (QTP) and Mongolia. Taxonomic ambiguity still exists for this sub-species because of complex evolutionary history anduse of limited wild samples across its range in Himalaya. We document for the first time population genetic structure and taxonomic affinity of the wolves across western and eastern Himalayan regions from samples collected from the wild (n = 19) using mitochondrial control region (225bp). We found two haplotypes in our data, one widely distributed in the Himalaya that was shared with QTP and the other confined to Himachal Pradesh and Uttarakhand in the western Himalaya, India. After combining our data withpublished sequences (n = 83), we observed 15 haplotypes. Some of these were shared among different locations from India to QTP and a few were private to geographic locations. A phylogenetic tree indicated that Woolly wolves from India, Nepal, QTP and Mongolia are basal to other wolves with shallow divergence (K2P; 0.000-0.044) and high bootstrap values. Demographic analyses based on mismatch distribution and Bayesian skyline plots (BSP) suggested a stable population over a long time (~million years) with signs of recent declines. Regional dominance of private haplotypes across its distribution range may indicate allopatric divergence. This may be due to differences in habitat characteristics, availability of different wild prey species and differential deglaciation within the range of the Woolly wolf during historic time. Presence of basal and shallow divergence within-clade along with unique ecological requirements and adaptation to hypoxia, the Woolly wolf of Himalaya, QTP, and Mongolian regions may be considered as a distinct an Evolutionary Significant Unit (ESU). Identifying management units (MUs) is needed within its distribution range using harmonized multiple genetic data for effective conservation planning.

Assessment and prediction of spatial patterns of human-elephant conflicts in changing land cover scenarios of a human-dominated landscape in North Bengal.

It is of utmost importance to research on the spatial patterns of human-wildlife conflicts to understand the underlying mechanism of such interactions, i.e. major land use changes and prominent ecological drivers. In the north eastern part of India there has been a disparity between nature, economic development and fragmentation of wildlife habitats leading to intense conflicts between humans and Asian elephants (Elephas maximus) in recent times. Both the elephant and human population have increased in the past few decades with large tracts of forests converted to commercial tea plantations, army camps and human settlements. We analyzed data maintained by the wildlife department on human deaths and injuries caused by elephant attacks between 2006-2016 to understand spatial and temporal patterns of human-elephant conflict, frequency and distribution. The average annual number of human deaths and injuries to elephant attacks between 2006 to 2016 was estimated to be 212 (SE 103) with the highest number of such incidents recorded in 2010-2011. Based on a grid based design of 5 km2 and 25 km2 resolution, the main spatial predictors of human-elephant conflicts identified through Maxent presence only models are annual mean precipitation, altitude, distance from protected area, area under forests, tea plantations and agriculture. Major land use changes were assessed for this region from 2008 to 2018 using satellite imageries in Arc GIS and a predicted imagery of 2028 was prepared using Idrisi Selva. Based on the 2018 imagery it was found that forest area had increased by 446 km2 within 10 years (2008-2018) and the annual rate of change was 12%. Area under agriculture had reduced by 128 km2 with an annual (-) rate of change of 2.5%. Area under tea plantation declined by 307 km2 with an annual (-) rate of change of 12% whereas area under human settlements increased by 61 km2 with an annual (-) rate of change of 44%. Hotspots of human-elephant conflicts were identified in an east west direction primarily around protected areas, tea plantations and along major riverine corridors. During informal interactions with farmers, tea estate labors it was revealed that local community members chased and harassed elephants from agriculture fields, human settlements under the influence of alcohol and thus were primary victims of fatal interactions. Our analytical approach can be replicated for other species in sites with similar issues of human-wildlife conflicts. The hotspot maps of conflict risk will help in developing appropriate mitigation strategies such as setting up early warning systems, restoration of wildlife corridors especially along dry river beds, using deterrents and barriers for vulnerable. Awareness about alcohol related incidents and basic biology of elephants should be organized regularly involving non-governmental organizations targeting the marginalized farmers and tea estate workers.

Understanding drivers of human-leopard conflicts in the Indian Himalayan region: Spatio-temporal patterns of conflicts and perception of local communities towards conserving large carnivores.

Human killing is the decisive and most critical expression of human-leopard conflict and needs to be addressed sensitively to maintain local support for leopard conservation in India. This research was undertaken to investigate the ecological aspects of human killing and injury, spatial characteristic and pattern of such sites, temporal and seasonal trends of attacks and perception of local communities towards leopard in the Indian Himalayan region (IHR). We surveyed two sites i) Pauri Garhwal in the western part and ii) North Bengal (Dooars) in the eastern part of IHR, compiled secondary data on human-leopard conflict records and made field visits to (N = 101) conflict sites. We also conducted (N = 186) semi-structured questionnaire surveys in each of the sites to assess perception of local communities towards leopard. We analyzed the conflict data using rare events model in a binary logistic regression framework to understand spatial patterns of such incidents for Pauri Garhwal and North Bengal. The average number of injuries and deaths to leopard attacks in Pauri was estimated to be 11 (SE 1.13) and 3 (SE 0.6) per year between 2006-2016 whereas in North Bengal it was estimated to be 70 (SE 9.2) and 1.6 (SE 0.3) respectively between 2004-2016. About 97% of the leopard attacks in North Bengal and 60% of the leopard attacks in Pauri resulted in human injuries. Majority of the leopard attack victims in Pauri were children and young people, whereas in North Bengal it was middle aged tea estate workers. Attack on humans in Pauri were recorded mostly near areas with dense scrub cover whereas in North Bengal it was reported within tea-estates. The percentage of human deaths to leopard attacks in Pauri were higher (40%) compared to a mere (3%) in North Bengal. Forty-one percent of respondents in Pauri and 75% in North Bengal were positive towards presence and conservation of leopard. A predictive risk map revealed central and northern regions of Pauri Garhwal and protected areas, peripheral areas in central and south-western dooars (North Bengal) as high "human-leopard conflict risk zones". This analytical procedure can be adopted in other sites to identify potential human-carnivore conflict risk zones.

Inertial microfluidics for sheath-less high-throughput flow cytometry.

Flow cytometer is a powerful single cell analysis tool that allows multi-parametric study of suspended cells. Most commercial flow cytometers available today are bulky, expensive instruments requiring high maintenance costs and specially trained personnel for operation. Hence, there is a need to develop a low cost, portable alternative that will aid in making this powerful research tool more accessible. In this paper we describe a sheath-less, on-chip flow cytometry system based on the principle of Dean coupled inertial microfluidics. The design takes advantage of the Dean drag and inertial lift forces acting on particles flowing through a spiral microchannel to focus them in 3-D at a single position across the microchannel cross-section. Unlike the previously reported micro-flow cytometers, the developed system relies entirely on the microchannel geometry for particle focusing, eliminating the need for complex microchannel designs and additional microfluidic plumbing associated with sheath-based techniques. In this work, a 10-loop spiral microchannel 100 microm wide and 50 microm high was used to focus 6 microm particles in 3-D. The focused particle stream was detected with a laser induced fluorescence (LIF) setup. The microfluidic system was shown to have a high throughput of 2,100 particles/sec. Finally, the viability of the developed technique for cell counting was demonstrated using SH-SY5Y neuroblastoma cells. The passive focusing principle and the planar nature of the described design will permit easy integration with existing lab-on-a-chip (LOC) systems.

Inertial microfluidics for continuous particle separation in spiral microchannels.

In this work we report on a simple inertial microfluidic device that achieves continuous multi-particle separation using the principle of Dean-coupled inertial migration in spiral microchannels. The dominant inertial forces coupled with the Dean rotational force due to the curvilinear microchannel geometry cause particles to occupy a single equilibrium position near the inner microchannel wall. The position at which particles equilibrate is dependent on the ratio of the inertial lift to Dean drag forces. Using this concept, we demonstrate, for the first time, a spiral lab-on-a-chip (LOC) for size-dependent focusing of particles at distinct equilibrium positions across the microchannel cross-section from a multi-particle mixture. The individual particle streams can be collected with an appropriately designed outlet system. To demonstrate this principle, a 5-loop Archimedean spiral microchannel with a fixed width of 500 microm and a height of 130 microm was used to simultaneously and continuously separate 10 microm, 15 microm, and 20 microm polystyrene particles. The device exhibited 90% separation efficiency. The versatility of the device was demonstrated by separating neuroblastoma and glioma cells with 80% efficiency and high relative viability (>90%). The achieved throughput of approximately 1 million cells/min is substantially higher than the sorting rates reported by other microscale sorting methods and is comparable to the rates obtained with commercial macroscale flow cytometry techniques. The simple planar structure and high throughput offered by this passive microfluidic approach make it attractive for LOC devices in biomedical and environmental applications.

Continuous particle separation in spiral microchannels using Dean flows and differential migration.

Microparticle separation and concentration based on size has become indispensable in many biomedical and environmental applications. In this paper we describe a passive microfluidic device with spiral microchannel geometry for complete separation of particles. The design takes advantage of the inertial lift and viscous drag forces acting on particles of various sizes to achieve differential migration, and hence separation, of microparticles. The dominant inertial forces and the Dean rotation force due to the spiral microchannel geometry cause the larger particles to occupy a single equilibrium position near the inner microchannel wall. The smaller particles migrate to the outer half of the channel under the influence of Dean forces resulting in the formation of two distinct particle streams which are collected in two separate outputs. This is the first demonstration that takes advantage of the dual role of Dean forces for focusing larger particles in a single equilibrium position and transposing the smaller particles from the inner half to the outer half of the microchannel cross-section. The 5-loop spiral microchannel 100 microm wide and 50 microm high was used to successfully demonstrate a complete separation of 7.32 microm and 1.9 microm particles at Dean number De = 0.47. Analytical analysis supporting the experiments and models is also presented. The simple planar structure of the separator offers simple fabrication and makes it ideal for integration with on-chip microfluidic systems, such as micro total analysis systems (muTAS) or lab-on-a-chip (LOC) for continuous filtration and separation applications.