Snakes represent a taxonomically underdeveloped group of animals in India with a lack of experts and incomplete taxonomic descriptions being the main deterrents to advances in this area. Molecular taxonomic approaches using DNA barcoding could aid in snake identification as well as studies of biodiversity. Here a non-invasive sampling method using DNA barcoding is tested using skin exuviates. Taxonomically authenticated samples were collected and tested for validation and comparisons to unknown snake exuviate samples. This approach was also used to construct the first comprehensive study targeting the snake species from Maharashtra state in India. A total of 92 skin exuviate samples were collected and tested for this study. Of these, 81 samples were successfully DNA barcoded and compared with unknown samples for assignment of taxonomic identity. Good quality DNA was obtained irrespective of age and quality of the exuviate material, and all unknown samples were successfully identified. A total of 23 species of snakes were identified, six of which were in the list of Endangered species (Red Data Book). Intra- and inter-specific distance values were also calculated, and these were sufficient to allow discrimination among species and between species without ambiguity in most cases. Two samples were suspected to represent cryptic species based on deep K2P divergence values (>3%), and one sample could be identified to the genus level only. Eleven samples failed to amplify COI sequences, suggesting the need for alternative PCR primer pairs. This study clearly documents how snake skin exuviates can be used for DNA barcoding, estimates of diversity and population genetic structuring in a noninvasive manner.

BackgroundThe hypothalamic nuclei constitute that part of the corticodiencephalic mechanism that activates, controls and integrates the peripheral, autonomic mechanisms, endocrine activity and many somatic functions. Their full integration into behavior requires the neocortex, in particular the limbic system.PurposeAn antiserum against histamine was used to reveal the location and time of appearance of the amine in developing fetal hypothalamus. Although the expression pattern of histamine is well studied in mammals, less is known about it in humans. The neurotransmitter histamine plays a crucial role in co-ordinating mutiple inputs from various brain centres.MethodsIn the present investigations the hypothalamus was studied in the human fetus ranging from 19 weeks of gestation (GW) to term by using immunocytochemistry. Distinct neurons that stained with Pischinger’s methylene blue were obsereved from 19 GW to term. Sections adjacent to those that contained neurons were stained for histamine labeling. Immunoreactive neurons in the hypothalamus at 19 GW of gestation show relatively meager population.ResultsHistamine immunoreactive (His-ir) neurons of the hypothalamus were divisible in lateral and ventrolateral subgroups at 19 GW to 24 GW At 32 GW along with the neuronal cell bodies some beaded fibers were visible. There was progressive increase in the histamine expressing neurons as the fetus grows. In term fetal specimens, two to three small groups of his-ir neurons gradually merged in a single large ventrolateral group.ConclusionThe findings of the present study provide for a better understanding of the chemoneuroarchitecture of histamine containing neurons in hypothalamus during second and third trimester of human fetal development.

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