Abstract

In forensic science, morphology and pollen grain identification are crucial as they provide useful data for cold case examination, crime scene analysis, and geographic tracking. The goal of the study is to create a trustworthy process for recognizing and classifying pollen grains according to their morphological characteristics, like size, shape, surface ornamentation. Using sterile tools like brushes, needles, or forceps, pollen is systematically extracted from flowers and prepared by mounting it on a glass slide using a mounting liquid like glycerin. The samples are then analyzed using light microscopy. Data analysis involves the measurement and summarization of pollen traits, using software packages for accurate computation and classifications. High-resolution images are examined to identify characteristic structural features, which enable the identification of specific plant species. The research seeks to advance the knowledge of pollen diversity and explain the impact of environmental and geographic conditions on its morphology and distribution. Expected results include the development of a standard methodology for forensic palynology, improved pollen grain classification, and the establishment of pollen as a major forensic tool for crime scene linkage to specific locations. The purpose of this research is to improve environmental forensics and forensic investigations.

Keywords: Palynology; Pollen grain; Morphology

Introduction

Introduction to Pollen Morphology

A type of study termed pollen morphology studies the nature, layout, and physical properties of pollen grains. Male seed plant gametophytes called pollen grains serve as a source of genetic information during fertilization. Forensic science, botany, paleontology, and allergy investigations all gain insight from the study of pollen morphology [1,2].
Dimensions and Form: The pollen grain sizes vary and are generally between 10 to 200 micrometers. The pollen grain shapes are spherical, oval, triangular, or irregular depending on the plant species.
Intine and Exine: It has two layers of pollen wall:
Exine: The highly resistant, external layer made up of sporopollenin. It has sculpturing patterns (reticulate, striate, etc.) that are characteristic of various plant species, making it extremely important for identification.

Intine is the inner coating made of cellulose and pectin, which is less resistant and thinner in nature.
Surface Decoration: The surface of the exine may exhibit different textures and ornamentations like striations, ridges, spines, or reticulations, characteristic of plant groups.
Forensic Palynology: Pollen analysis helps link suspects to crime scenes by identifying the precise vegetation in an area.

1. Size:

•The pollen grain is between 5 μm (e.g., Myosotis) and over 200 μm (e.g., Cucurbitaceae).
•on the equatorial diameter or the polar axis.

2. Shape:

It is defined by the ratio of the polar axis (PA) to the equatorial diameter. (ED):
•Spheroidal (PA ≈ ED).
•Prolate (PA > ED).
•Oblate (PA < ED).

3. Symmetry:

Radiosymmetric: Having symmetry in any plane passing through the polar axis.
• Bilateral: Symmetry in one plane.
• Asymmetric: Not symmetrical or irregular.

4. Polarity:

Polar Pollen: Separate proximal and distal poles. • Apolar Pollen: No established poles.

5. Apertures:

Germination pores on the exine • Colpate: Furrows.
• Porate: Circular pores.
• Colporate: Furrows containing pores.
• Inaperturate: Apertureless.

6. Exine Ornamentation:

The external surface structure of the exine:
• Reticulate: Net-like.
• Spinate: Possessing spines.
• Granulate: With granules.
• Smooth: Plain surface.

7. Color and Transparency:

Depends on the composition of the exine and is observed in light Microscopy.
• Light Microscopy (LM): Initial evaluation of configuration, size, and symmetry.
• Scanning Electron Microscopy (SEM): Close study of surface ornamentations and Apertures.
• DNA Barcoding: Species identification at the molecular level.
• Automated Identification: AI-powered systems for fast classification.

It links suspects, victims, or objects to places by comparing unique or localized pollen profiles. It determines the time or season of a crime, as the pollen pattern typically follows seasonal patterns. It supports geographical profiling, especially in rural or ecologically distinctive areas. Provide assistance in cold cases or re investigation of the scene where other physical evidence is not sufficient. The goal is to examine and document the morphological characteristics of 50 plant species’ pollen grains through light microscopy with the view to creating a reference system for botanical identification and forensic use. To carry out this study, I picked the pollen grains from flowers directly using clean and precise equipment like a brush, needle, or forceps. This equipment are used to avoid contamination and hence pick only the intended pollen grains. The picking is done in the laboratory setting to ensure no contact with any external contaminants or mixed pollen from other organisms. Pollen should be picked carefully from different species of plants to create a diversified dataset that will be capable of capturing a wide range of pollen types in the environment. Once the collection process is complete, the pollen is transferred to a sterilized glass slide [3,4].

A portion of the prepared pollen is carefully put on the slide to avoid any type of distortion or damage. Glycerin or other mounting medium is used to protect the grains from further inspection to speed up their examination. By preventing drying out and retaining the pollen’s structure, glycerin acts as a protective coating that facilitates microscopic observation. The sample is covered in a coverslip after the mounting media is applied to prevent air bubbles from disrupting with the microscope image. The prepared slide is next viewed under a light microscope, firstly at low magnification (10x to 40x) then at higher power (100x). At low magnification, pollen grains are usually easier to find and where they are on the slide can be determined. At greater magnification levels, the pollen grains’ finer structural characteristics-such as their size, shape, surface roughness, and the existence of apertures-more particularly, holes or pores--can be observed. These unique characteristics are crucial for distinguishing between plant species because each species produces pollen grains that have distinctive structural characteristics.

Under microscopic examination, the different morphological characters of the pollen grains are observed. The most important observations to be recorded are as follows:
• Shape: The grain’s general shape, which may be spherical, oblate, or triangular.
• Dimensions: The pollen grains’ dimensions, typically stated in micrometres.
• Surface Decoration: A detailed analysis of the surface motifs, including texture, reticulation, spines, or smoothness, typical for each plant Species. High-resolution photographs are obtained at various magnifications, allowing for detailed documentation with respect to pollen morphology. The closeup photos are saved for subsequent analysis and Classification. Once they are collected and recorded, the data are processed using sophisticated software tools. These instruments enable the measurement and integration of the major characteristics of the pollen grains, such as dimensions, configuration, aperture classification, and surface embellishments. Computer software may calculate the the sizes of pollen grains and classify them into some categories depending on their Morphological features. A comparative analysis can be conducted to establish patterns and similarities between various species, and any differences that may be attributed to Environmental or geographical determinants. In addition, high-resolution images taken using the light microscope are subjected to identify any distinctive structural features of the pollen grains. This careful assessment will be beneficial The recognition and categorization of various pollen types are significant for Establishing relationships between pollen records and individual geographical sites or plant types (Table 1).

A total of 50 pollen samples from various angiosperms were studied. Morphological parameters observed: pollen shape, size and surface ornamentation. The most common shape found here was spheroidal. The size was varied from 10 - 100μm. By documenting pollen morphology from 50 species, this study is a useful addition to the development of reference databases relevant to botany, ecology, and forensic science. The morphological features documented can be useful to forensic investigation by revealing the geographic origin of materials or linking individuals to crime scenes through pollen evidence. Pollen grains are extremely resilient and have the capacity to hold on to clothing, vehicles, equipment, and human bodies. The morphological description of the 50 species serves as a reference for the identification and linking of pollen traces on suspects or at crime scenes.

Most plant species are endemic to specific geographic regions. The identification of various types of pollen can be used to link a suspect or object to a particular location or environment, thus limiting the possibilities of the crime scene. Comparison with Punt et al. [5,6]. Punt’s pollen type classification system, in terms of their apertures and ornamentation, is repeated in this study. These classifications in morphology are utilized in this research and confirm their effectiveness in separating species under light microscopy. Comparison with Horrocks & Walsh [7-10]. That study pointed out the value of pollen in forensic science, especially in geographical profiling. Similarly, this study points out the forensic value of pollen, but with an expanded dataset (50 species), thus offering a larger reference source for forensic palynologists. The detailed morphological features of pollen grains, such as nanoscale ornamentation and complex surface patterns, tend not to be easily visible, and this contributes to limitations in the precise identification of their morphology. Only 50 pollen samples were examined, and these might not be representative of the overall morphological variability of pollen from all plant groups and ecological communities. Most pollen grains of taxonomically related genera or species are of comparable size and shape, and species-level identification is ambiguous in the lack of molecular data.

Conclusion

The morphological study and identification of pollen is essential in forensic science, ecology, plant taxonomy and other areas. Due to their unique shape and size, their surface features, and ornamentation, pollen grains enable the identification of various planets, from which different species, is found. Of all types of forensic traces, pollen grains are the most fragile. This is because not only are they small, but they also possess a wide range within a species and are usually preserved very well. In forensic botany, the movement and location of subjects, victims, and objects within a crime scene can be easily identified using pollen analysis. Pollen, being minute in size, can easily stick on garments, hair, property, and other items, creating an important link between a person, the crime scene, and accompaniment. For instance, the occurrence of the rare pollen of a certain plant with limited geographical distribution can prove that a certain area was visited by a suspect. In addition to establishing or tracing the source of certain materials, seasonality, or even undisclosed graves can be determined. With the integration of palynology in forensic examination, the credibility and dependability of sceneries in the crime scene reconstruction is received high precision. It enables a more accurate visualization and understanding of the series of events and scenario surrounding a specific area of interest. This study affirms that pollen morphology and identification enhance the capabilities of forensic science and enable it to realize the fulfilment of justice.

Acknowledgement

I would like to thank Mrs. Girija Kumari R, who offered laboratory facilities and support throughout this study. Special thanks are due to those who offered helpful suggestions, continuous encouragement, and constructive criticisms throughout the study. I would like to thank the team and colleagues who assisted in the identification and collection of plant specimens. I also acknowledge the invaluable contribution of the microscopy equipment and other research equipment provided by the Lifecare Clinical Laboratory.

References

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