Forensic Palynology Research Articles

Pollen morphology of Asteraceae from Garhwal Himalaya (Uttarakhand, India): part I

Palynotaxonomy is crucial for understanding plant evolution, biodiversity, and ecological history by systematically classifying and identifying species based on pollen morphology. In addition, a regional pollen flora is crucial for precise pollen identification in various applied fields of palynology, including melissopalynology, aeropalynology, forensic palynology, paleopalynology, and copropalynology. In this communication, we explore the pollen morphology of 28 species of Asteraceae, employing both light (LM) and scanning electron microscopy (SEM). The results revealed that the grains were monad, isopolar, radially symmetric, tricolporate, and exhibited oblate-spheroidal to prolate-spheroidal shapes. The infratectum was found to be columellate, caveate, or noncaveate. The supratectum exhibited echinate or microechinate characteristics, with the exine surface showing perforate, microreticulate, or perforate regulate features. Among some species, a lophate pattern was observed, revealing two distinct variations: one characterized by a ‘15 lacunae type,’ while the other exhibited a ‘21 lacunae type’. Some unusual variations were observed in the aperture character among certain species, like the occurrence of tricolpate grains and pollen dimorphism due to variation in aperture number. Light microscopic examination was preliminary for resolving columellar structure but it clearly showed a double level of columella in some species. A cluster analysis, incorporating qualitative and quantitative features of pollen grains and an artificial pollen key have been set forth to distinguish studied species, supplemented by LM and SEM micrographs. The current findings have enriched the existing pollen database of Asteraceae and could play an important role not only in palynotaxonomy but also in allied fields.

Pollen morphology of Acanthoideae species from the Shivalik Foothills in Rajaji National Park (Uttarakhand, India).

Apart from its role in plant taxonomy, the understanding of pollen morphology is considered an essential interdisciplinary tool in diverse applied fields, including plant systematics, melissopalynology, aeropalynology, forensic palynology, paleopalynology, and copropalynology. In palynotaxonomy, it is frequently employed to classify and validate natural groups across a range of hierarchical levels, from higher categories down to the infrageneric level. The subfamily Acanthoideae, within the Acanthaceae family, consists of a diverse group of flowering plants that are distributed globally. The present study attempted a comprehensive analysis of the pollen morphology, employing both light microscopy (LM) and scanning electron microscopy (SEM), for a total of 13 Acanthoideae species from the Shivalik Foothills in Rajaji National Park (located in the northern Indian state of Uttarakhand, Western Himalaya). The findings indicated that the Acanthoideae is characterized by eurypalynous features, and the studied species exhibited pollen grains that were monads, radially symmetrical, and varied in size from small to large. The pollen grains were predominantly tricolporate or heteroaperturate, with porate occurrences being rare. The significant variation in exine sculpturing, including reticulate, coarsely reticulate, and bireticulate patterns, holds substantial taxonomic significance. The detailed presentation encompasses pollen morphological characters described with LM and SEM micrographs, along with a species-level identification key. RESEARCH HIGHLIGHTS: This article provides a thorough analysis of the pollen morphology of Acanthoideae taxa using both light microscopy and scanning electron microscopy, covering 13 species across 10 genera and revealing a broad spectrum of pollen characteristics, including size, shape, aperture type, and exine sculpturing. The microscopic investigation of these Acanthoideae species not only enhances our understanding of their pollen morphology but also aids in species identification through the development of a pollen-based key.

Stepping towards pollen DNA metabarcoding: A breakthrough in forensic sciences.

This review is engaged in determining the capability of plant pollen as a significant source of evidence for the linkage between suspects and crime location in forensic sciences. Research and review articles were collected from Google Scholar, the Web of Science, and PubMed. Articles were searched using specific keywords such as "Forensic Palynology," "Pollen metabarcoding," "Plant forensics," and "Pollen" AND "criminal investigation." Boolean logic was also utilized to narrow the articles to be included in this review article. Through the literature and exploratory research, it has been observed in the current study that with advancements in technology, forensic palynology has found its application in creating an association between the crime scene and suspected individuals to have a link to it, as pollen DNA is a long-lasting investigative tool that can effectively help forensic investigations. Moreover, the literature shows that the DNA of pollen and spores has helped forensic scientists link suspects to crime scenes, and the introduction of pollen DNA metabarcoding tools has eased the efforts of palynologists to analyze pollen DNA. The introduction of DNA metabarcoding techniques to analyze pollen from plants has helped identify the geological locations of the plants and ultimately identify the culprit.

Use of pollen assemblages as forensic evidence in non-seasonal high-altitude soils

Forensic palynology is a tool in criminalistics that uses spores and pollen grains to link a certain geographical location with a crime scene. The comparison of the pollen assemblage of a crime-scene soil and that of footwear of suspects and victims proved to be very useful as judicial evidence in multiple environments with marked seasonality. However, its usefulness in non-seasonal high-altitude soils has not been experimentally evaluated to the same extent. For this reason, the present study addressed this information gap by undertaking a palynological study in areas with high crime rates in the city of La Paz, Bolivia. To do this, we carried out multiple experimental samplings in three locations with different types of soil and different degrees of urbanization. Specifically, we compared whether the vegetation present at the time of taking the reference samples, was reflected in the pollen rain. Results showed that the vast majority of the species present in the vegetation were found in the pollen rain, with the exception of some plant species with entomophilous pollination syndrome. We also show that the transfer between assemblages from pollen rain to footwear happened effectively, which helped identify their geographical origin, and unveiled a great number of useful indicator species.

The application and benefits of forensic palynology in robbery events in Turkey

Forensic science is required to elucidate crime, and it has been carried out through various induction methods. Forensic palynology, a branch of forensic science, is used to find offenders by investigating the connection between the crime scene, evidence, and offender, solving events, proving the reliability of the victim's narratives, building up the profile of the offender, reducing the number of suspects, assisting security forces in conducting accurate investigations and detecting smuggling of drugs and other materials. Palynological evidence is known to be extremely resistant to extremely high temperatures (over 400°C), strong acids (such as HCl-HF), and fungal and bacterial activity. One of the most important reasons for using palynological evidence as evidence in court is its high resistance. Property crimes account for most judicial cases in Turkey. Burglary accounts for a significant portion of property crime. We aimed to solve various burglary events in Turkey using palynological evidence in this study. We used various methods to analyze palynological particles found in forensic samples sent to our laboratories between March 2016 and May 2017. The findings were distributed to law enforcement officers and legal experts working in courtrooms (lawyers/attorneys/judges).

Forensic potential of pollen grains: A case study from continental Croatia (town of Donja Stubica)

Background and Purpose: Pollen grains can come as “ invisible” traces during forensic palynological expertise. As forensic palynology is not carried out in Croatia at all, the main aims of our work were to establish an initial palynological collection for a limited area in Croatia and determine its forensic potential. Materials and Methods: For our case study, we chose a typical town from continental Croatia, Donja Stubica, in which there had been no previous floristic studies. The field survey was conducted during 2005, 2007, and 2021. Floristic and palynological analysis was carried out with an emphasis on the forensic potential of pollen grains. Results: A total of 141 representative plant taxa with pollen samples were collected. The taxonomic analysis showed that the area for the case study was chosen well. Palynological analysis showed that the highest number of taxa had pollen grains that were spheroidal in shape (60%), medium sized (70%), and colporate (41%). The pollen of the majority of taxa had excellent or very important forensic potential, due to their dispersal by insects (60%), a combination of two dispersal methods (23%), and autogamy (1%). In addition, due to the limited distribution of certain rare plants (e.g., Iris sibirica subsp. sibirica in Croatia), their pollen has an even greater forensic potential than the similar pollen of related, more widespread taxa. Conclusion: The generated palynological collection, due to the great forensic potential, could be used in the future as a comparative, and at least partly as a reference forensic collection, applicable in specific cases of legal pollen expertise in Croatia.

Palynological Evidence from Sediment Samples Associated with the Early Holocene Human Skeleton from Fa Hien-lena, Sri Lanka

This paper highlights the value of pollen-analytical studies in the prehistoric archaeological context at the Fa Hien rock shelter. The Fa Hien rock shelter located in the Wet Zone of southwestern Sri Lanka has been occupied by the earliest anatomically modern humans in South Asia, dating from the late Pleistocene to the middle Holocene. The pollen evidence from the samples associated with the early Holocene human skeleton from this site has been used to assess the burial context and palynological taphonomy. Pollen data analysis indicates that high taphonomic impact occurred on the pollen grains in the depositional context, possibly due to reworking, burning, mechanical pressure and other possible human and animal activities. Furthermore, the pollen assemblage appears to have been primarily derived from the lowland rainforest, including disturbed habitats. Along with archaeological stratigraphy and radiometric dating, the pollen assemblage can be used to make broad inferences on the burial contexts and environment of the rock shelter occupants, while discussing the significance of forensic palynology in the Fa Hien archaeological context.

Forensic palynology: computer vision and geotechnologies to support criminalistics expertise

Pollen grains can provide valuable information to forensic palynology, such as the time of death or the possible origin of a corpse. Forensic Palynology is a vital tool to be used in a criminal investigation because the different environment has distinct pollen signatures. Brazil has a rich and diversified flora that is suitable for the application of forensic palynology. The purpose of this research is to introduce palynology automation as a tool to improve the investigative method in forensic palynology and apply it to forensic palynology automation. The studied city has different vegetation types, in which we performed assessments to identify its correspondent pollen types. PALINOVIC algorithm was developed using computer vision and geotechnology techniques. Our results show that it is possible to correlate pollen grains found in forensic samples by automatic pollen identification and with a mapping of the likely vegetation. Our results show that it is possible to relate the presence of pollen grains found in forensic samples through the automatic identification of images together with a database of georeferenced plant species. It was possible to analyze the pollen grains collected in eight bodies, where the algorithm presented a performance of 90.51% in the pollen grain classification tests. Furthermore, pollen grains could be correlated with the type of vegetation where the body was found. Thus, the technique developed can be applied in other urban centers from a previous georeferencing of plants, as well as a pollen database.

Forensic entomology: A novel approach in crime investigation

Forensic entomology is an application of studies related to insects and other arthropods in legal matters. Depth knowledge and experience of the biodiversity, biology, ecology, and behavior of forensic insects found on a human corpse or at a crime site, can provide information about time since death, location of the crime, and in some cases cause of death. Although forensic insects are used in crime investigation for three to four decades, it aroused recently as a very popular and independent branch of forensic science in crime investigation. While observing and examining insects on a corpse, knowledge of regional insects’ assemblages and their population dynamics play an important role in medico-legal entomology for an efficient criminal investigation. Forensic entomology has three components, named as urban, stored products, and medico-legal. As different groups of insects are attracted to different stages of decay, forensic insects invade the corpse in a typical sequence. The type of insects’ colonization found on corpses provides important information about the decomposition stage. Calliphoridae provides valuable evidences, particularly in determining time since death in criminal matters, where the time since death is prolonged and use of other conventional methods for estimation of time since death is limited. Two well-established ways used for estimation of post mortem interval are based on maggot’s development and succession waves of entomofauna. The latest scientific and technological developments in the field, such as its combined approach with forensic toxicology, forensic palynology, DNA based molecular identification of insect species, and other various methods to estimate post mortem interval, are many emerging fields under this periphery and forensic entomology lately became very popular among criminal investigators and forensic entomologists.

CURRENT TRENDS IN FORENSIC PALYNOLOGY: A REVIEW STUDY

Forensic Palynology is the study of natural organic matter to establish a chronological or environmental history. It is often used in palynology (palynology being the study of pollen and spores) to reconstruct past climate and vegetation changes. If the stratigraphic setting is known, the deposition date can be determined by organic matter in the sediment. The primary factors which dictate the abundance of organic matter are changes in climate and vegetation. Modern forensic palynology has made considerable advances in examining environmental samples, with many studies now showing that the "structure of pollen grains, in particular, provides an indicator of a sample's environmental and occupational history." It, therefore, provides "a way of finding out what happened before and during an environmental exposure." The field of forensic palynology is concerned with the study and application of plant and soil organic and biotic evidence objectively and scientifically in order to provide evidence of environmental and occupational processes and conditions".

Current Trends in Forensic Palynology

Forensic Palynology is the study of natural organic matter to establish a chronological or environmental history. It is often used in palynology (palynology being the study of pollen and spores) to reconstruct past climate and vegetation changes. If the stratigraphic setting is known, the deposition date can be determined by organic matter in the sediment. The primary factors which dictate the abundance of organic matter are changes in climate and vegetation. Modern forensic palynology has made considerable advances in examining environmental samples, with many studies now showing that the "structure of pollen grains, in particular, provides an indicator of a sample's environmental and occupational history." It, therefore, provides "a way of finding out what happened before and during an environmental exposure." The field of forensic palynology concerned with the study and application of plant and soil organic and biotic evidence objectively and scientifically in order to provide evidence of environmental and occupational processes and conditions".

Network-based geoforensics: Connecting pollen and plants to place

Despite the potential for pollen to provide key geographic information, law enforcement agencies have historically underutilized forensic palynology. Pollen samples are rarely collected at crime scenes or on objects of interest, and experts needed to manually identify pollen and its morphological characteristics are often in short supply. Fortunately, the advent of DNA barcoding and metabarcoding is poised to change this, enabling law enforcement agencies to easily and quickly identify individual plant species from mixed pollen samples. However, determining the location of pollen deposition based on identified plant species remains challenging, requiring comprehensive plant geodatabases and complex analytical approaches. This type of analysis is also complicated because pollen samples from forensic objects are not comprehensive. Furthermore, while detecting common plant pollen in a sample is likely, common plants provide less explanatory power due to their ubiquity. Thus, we propose a network-based approach to construct a categorization of ‘forensically-useful’ plants – those species that characterize geographically distinct plant mixes quickly for forensic applications and thus provide significant explanatory power to a geolocation model. The developed tool can better inform more detailed forensic search models, significantly reducing their overall computational burden and enabling high-fidelity forensic computational tools to operate faster, at a higher resolution, and over a more extensive study area.

Identifying the spatial footprint of pollen distributions using the Geoforensic Interdiction (GOFIND) model

Geoforensics leverages various spatial analytical, biological, ecological, and geological techniques to improve criminal investigations, including efforts to thwart terrorism, evaluate humanitarian crises or identify the origin of fraudulent goods. In addition to investigating what took place, geoforensic efforts often focus on determining where and when the event occurred. Forensic palynology is a promising subfield of geoforensics that uses pollen to link persons or objects to particular places and times. This promise is driven by pollen's strong utility as a biomarker. It is ubiquitous, durable and demonstrates a relatively predictable distribution in time and space. As a result, pollen often provides important clues for determining the provenance of hard-to-trace items, such as computers, fraudulent goods, digging tools, clothing, and undetonated explosives during geoforensic investigations. One major limitation that has reduced the implementation of forensic palynology is the lack of computational tools that relate pollen species to geographic locations. The purpose of this paper is to introduce a robust geocomputational framework that uses pollen sample composition to identify the relative likelihood of potential origin locations for improving geoforensic efforts. Using USDA CropScape data for the state of Texas, our results suggest that this framework allowed for multiple potential origin sites to be identified simultaneously, with solution properties that were better than a random process and offer a possible alternative to the single site joint probability approaches commonly used in the field of forensic palynology.

Grass Gazers: Using citizen science as a tool to facilitate practical and online science learning for secondary school students during the COVID-19 lockdown.

The coronavirus disease of 2019 (COVID‐19) pandemic has impacted educational systems worldwide during 2020, including primary and secondary schooling. To enable students of a local secondary school in Brisbane, Queensland, to continue with their practical agricultural science learning and facilitate online learning, a “Grass Gazers” citizen science scoping project was designed and rapidly implemented as a collaboration between the school and a multidisciplinary university research group focused on pollen allergy. Here, we reflect on the process of developing and implementing this project from the perspective of the school and the university. A learning package including modules on pollen identification, tracking grass species, measuring field greenness, using a citizen science data entry platform, forensic palynology, as well as video guides, risk assessment and feedback forms were generated. Junior agriculture science students participated in the learning via online lessons and independent data collection in their own local neighborhood and/or school grounds situated within urban environments. The university research group and school coordinator, operating in their own distributed work environments, had to develop, source, adopt, and/or adapt material rapidly to meet the unique requirements of the project. The experience allowed two‐way knowledge exchange between the secondary and tertiary education sectors. Participating students were introduced to real‐world research and were able to engage in outdoor learning during a time when online, indoor, desk‐based learning dominated their studies. The unique context of restrictions imposed by the social isolation policies, as well as government Public Health and Department of Education directives, allowed the team to respond by adapting teaching and research activity to develop and trial learning modules and citizen science tools. The project provided a focus to motivate and connect teachers, academic staff, and school students during a difficult circumstance. Extension of this citizen project for the purposes of research and secondary school learning has the potential to offer ongoing benefits for grassland ecology data acquisition and student exposure to real‐world science.

POLLEN73S: An image dataset for pollen grains classification

The classification of pollen species and types is an important task in many areas like forensic palynology, archaeological palynology, and melissopalynology. This paper presents a new public annotated image dataset for the Brazilian Savanna called POLLEN73S composed of 2523 images from 73 pollen types. Using the state-of-the-art Convolutional Neural Networks (CNNs), we provide a baseline for pollen grain classification. Our experiments showed evidence that DenseNet-201 and ResNet-50 have superior performance against the other CNNs tested, achieving precision results of 95.7% and 94.0%, respectively. Due to its category coverage and satisfactory diversity of examples, POLLEN73S offers a diversity of pollen grain to guide progress in computer vision to solve Palynology problems.

Non-destructive DNA analysis of single pollen grains

Forensic palynology has played a crucial role in many criminal investigations, from burglary to homicide, rape, war crimes, terrorism, and drug trafficking. With the emergence of DNA barcoding, pollen genomics has immense potential to classify and identify pollen taxonomically from its genetic signature. In forensic analysis, a non-destructive method for evidence is essential. Thus, several challenges exist for the extraction of DNA from the pollen grains, such as keeping the grain intact for secondary analysis and bypassing the issue of DNA mixtures more likely to be observed in a pollen assemblage. In this study, a new method was developed to address these challenges and was designed to extract genetic material while not destroying the pollen grain on a single grain level. We demonstrate the mehod's efficiency with Pinus echinata, Taxodium distichum, and Plantago lanceolata single pollen grains. Quantitative PCR amplification of two genetic markers, ribulose bisphosphate carboxylase (rbcL) and internal transcribed spacer 2 (ITS2), and digital microscopy demonstrates the non-destructive nature of this novel DNA extraction method from single pollen grains.

Research Progress on Forensic Palynology and Its Application in Forensic Science.

Forensic palynology is to apply palynology to the field of forensic science, using pollen and spores to solve issues in juridical practice, such as civil and criminal issues. Sporopollens have a small size, wide distribution, diverse morphology, can be easily transferred, have durability, and is not easily noticed. It can provide strong investigation and related evidence for case detection as good trace evidence. The application of palynology in forensic science has achieved certain success, but it is underutilized in most countries. This paper analyzes the evidence value provided by sporopollen, collection of the sporopollen samples, the progress in detection technology and challenges ahead, based on the biological characteristics of sporopollen, combined with recent successful cases in forensic science, to comprehensively discuss the research progress in forensic palynology and its application prospects in forensic science.

Pollen molecular biology: Applications in the forensic palynology and future prospects: A review.

Palynology, which is the study of pollen and spores in an archaeological or geological context, has become a well-established research tool leading to many significant scientific developments. The term palynomorph includes pollen of spermatophytes, spores of fungi, ferns, and bryophytes, as well as other organic-walled microfossils, such as dinoflagellates and acritarches. Advances in plant genomics have had a high impact on the field of forensic botany. Forensic palynology has also been used and applied more recently to criminal investigation in a meaningful way. However, the use of pollen DNA profiling in forensic investigations has yet to be applied. There were earlier uses of dust traces in some forensic analyses that considered pollen as a type of botanical dust debris. Pollen grains can be studied for comparative morphological data, clues to unexpected aspects relating to breeding systems, pollination biology and hybridization. This can provide a better understanding of the entire biology of the group under investigation. Forensic palynology refers to the use of pollen and other spores when it is used as evidence in legal cases to resolve criminal issues by proving or disproving relationships between people and crime scenes. This overview describes the various contributions and the significance of palynology, its applications, different recent approaches and how it could be further employed in solving criminal investigations.

Forensic palynology and the search for geolocation: Factors for analysis and the Baby Doe case

First used over 50 years ago, forensic palynology is an important tool for law enforcement agencies. In most countries that use forensic palynology, microscopic pollen grains and spores are traditionally used in criminal investigations to link suspects to crime scenes or items. While still underutilized in many parts of the world, forensic palynology is increasingly being used to determine the region of origin, or geolocation, for persons and items of interest. Drawing upon the experience of the authors using trace pollen and spores to geolocate forensic samples, the types, methods, and variables of this type of analysis are discussed and demonstrated using the Baby Doe case from Massachusetts, USA as a case study. This is not an exhaustive list and every forensic sample is unique so the methods and experience presented here are intended to be a guide for future forensic and anti-terrorism investigations as forensic palynology becomes more commonplace in law enforcement agencies around the world.

ABOUT APPLICATION POSSIBILITY OF POLLEN AND SPORE ANALYSIS FOR DETECTION OF AN AFFILIATION OF "SPACE" PLANT MATRIX FOR A SINGLE SOURCE OF ORIGIN

While criminal investigations concerning distribution and use of narcotic it is necessary to carry out research on plant matrices that are impregnated with a solution of a narcotic substance. At the same time, the issue of attribution to one batch of psychoactive substances impregnated to inactive matrixes of plant origin is of particular relevance at present. The research purpose is to study possibility of using a method of spore and pollen analysis to prove belonging of a series of samples of crushed plant material of various herbs: Bidens (Herba Bidentis), Hypericum (Herba Hyperici), foliage of Salvia (Salviae folia), foliage of Tussilago (Tussilaginis farfarae folia), Chamomile flowers (Matricariae flores) and the vegetative collection of "Stomaran" to a single source of origin. To achieve desired goal it was necessary to solve the following tasks: to determine whether there was a small sample of pollen and plant spores in a quantity sufficient to calculate the reliable percentage content of separate taxa; perform spore and pollen analysis of vegetative raw materials of several series of samples: a) belonging to one lot; b) from lots of different manufacturers; c) belonging to different lots of the same manufacturer; to reveal indicators testifying the belonging of comparable series of samples to single or different sources of origin. The results of the research indicate that the revealed features of the composition of the palinoflora and the percentage content of separate components of the spore and pollen spectra, obtained as a result of the spore and pollen analysis of plant material, allows to distinguish a number of important indicators on the basis of which a confident conclusion can be made about belonging to a single or various sources of origin. Consequently, it is possible to conclude that data of the spore and pollen analysis contains extremely important evidence and have undeniable practical significance for solving the actual expert and criminalistic tasks, and the use of this method is the beginning of a new trend in the forensic palynology in the Republic of Belarus.