eChapters

Site of incision: Detach the limb from the trunk by cutting various muscles viz; pectoral muscles, serratus ventralis, latissimus dorsi, brachiocephalicus, omotransversarius, trapezius, rhomboideus and long head of triceps.

Site of incision

Remove the skin flap of pectoral region to expose the pectoral muscles.

Site of incision: Make a straight dorso-ventral incision through skin from the summit of dorsal vertebral spine to the cranioventral aspect of shoulder joint. Extend it up to the cranial margin of shoulder joint, then caudo-ventrally up to caudal most part of elbow joint. Make another ventrodorsal straight incision from caudal most point of elbow joint to the summit of vertebral spine then reflect the skin flap from ventral to dorsal up to mid dorsal line of the body. Remove the facia to expose the various muscles of this region.

Myology: It is the branch of biological science which deals with the description of different muscles and their accessory structures.

Muscle is a contractile tissue present in all animals. Its function is to produce motion. Impulses from nerve cells control the contraction of each muscle fiber. Contractile part of muscles is muscular tissue.

There are three kinds of muscular tissue in the body:

a. Striated or striped muscle

b. Non-striated or unstriped or smooth muscle

c. Cardiac muscle.

For dissection purpose only striated muscles are included in this book.

Omics technologies based on genes and proteins involved in the nematode activities and the plant resistance responses are key factors to novel nematode management. Techniques such as next-generation sequencing of genomes and transcriptomes, RNAi, PIs, MAS, chemo-disruptive peptides and genetic transformation systems with reproducible results must be available within phytonematodes management strategies to enhance crop yields (Abd-Elgawad, 2022). Linking long-read sequencing to the use of high density genetic mapping can also support the detection and characterization of nematode-virulent genes. Progress in computational biology, bioinformatics, and analyzing the omics large-scale data can efficiently boost these techniques to offer accurate recognition of components and pathways engaged in nematode parasitism and plant response. Updated genome-editing devices will serve classical plant breeding and precisely translate how gene actions are linked to phenotypic performances. Yet, these methods must be cautiously used to avoid unwanted effects such as nematode virulence and pleiotropic impacts on qualitative and quantitative crop yields. Definite issues such as those related to the transfer of multiple disease resistance traits, gene original construct, and specificity of the virulent nematodes to the R-gene may arise in particular cases.

Modeling phytonematodes is helpful in the investigation of several host plant interactions and may yield useful information to be exploited in the integration of management strategies and biological control. Modeling the phytonematodes density changes in a soil/root microcosm may allow the identification of main factors active in such systems (Ciancio et?al?, 2022). The multiple tri-trophic interactions linking roots, phytonematodes and antagonists, however, originate a very complex system. It requires knowledge about the many parameters describing the nematode life stages, the interactions with the antagonistic organisms present in the microcosm, as well as their capabilities for effective and durable host regulation. Descriptive variables should also account for the effects of roots and of other environmental factors at work in the tri-trophic interaction system. Models may provide benefits, including the possibility of identifying the best moment for the inoculation of a biocontrol agent, the optimal amount of its inoculum applied to maximize its biocontrol efficacy, as well as the possibility to simulate and thus investigate its behavior once introduced in soil.

Genetic resistance is the more efficient and the more environmentally friendly way to protect crops against phytonematodes. Nematode management is complex, hence, the prevention of an initial phytonematode infestation in pathogen-free areas is of fundamental importance. However, after the entry of the nematode into an area, it is virtually impossible to eradicate because it is a soil-dwelling organism. In this regard, the genetic resistance of plants to nematodes is considered one of the most efficient and economically feasible methods to prevent production losses (Matsuo et?al?, 2012). Thus, breeding for nematode resistance is essential in crop management to obtain a high yield and stable production.

14.1 Model Organism: A model organism is a species that has been widely studied, usually because it is easy to maintain and breed in a laboratory setting and has particular experimental advantages. Model organisms are non-human species that are used in the laboratory to help scientists understand biological processes. They are usually organisms that are easy to maintain and breed in a laboratory setting. For example, they may have particularly robust embryos that are easily studied and manipulated in the lab, this is useful for scientists studying development. They may occupy a pivotal position in the evolutionary tree, this is useful for scientists studying evolution.

In the soil environment plants are constantly exposed to a range of microorganisms which are likely to influence one another, as they occupy the same habitat. Phytonematodes are often considered as pathogens in their own right and are capable of producing a single, recognizable disease. Apart from this, they also get associated with other soil pathogens that result in the complex diseases, which are more devastating and cause huge crop losses. Phytonematodes are major predisposing factors for other potential soil pathogens, which deserve more attention. In the case of soil borne pathogens, further opportunities exist for interactions with other microorganisms occupying the same ecological niche. The significant role of nematodes in the development of diseases caused by soil borne pathogens has been demonstrated in many crops throughout the world. In many cases, such nematode–fungus disease complexes involve root-knot nematodes (Meloidogyne spp.), although several other endoparasitic (Globodera spp., Heterodera spp., Rotylenchulus spp., Pratylenchus \spp.) and ectoparasitic (Xiphinema spp., Longidorus spp.) nematodes have been associated with diseases caused by soil borne fungal pathogens (Back et al., 2002).

Our understanding of plant–nematode interactions has increased significantly. The f irst genome sequences of two root-knot nematodes species, M? incognita and M? hapla, have been described, which were significantly different from genome of the free-living nematode, Caenorhabditis elegans. Both M? incognita and M? hapla have definite set of proteins that determine the virulence in plant species. The secretomes (set of secreted proteins through the stylets) of different phytonematodes have demonstrated a number of effector proteins that are involved in compatible plant nematode interactions (Ali et?al., 2017). In response to infection of various nematodes, plant’s transcriptome resulted in increased metabolic activity in the feeding cells and suppression of defense mechanisms of the plants in most of the cases. Most of these studies revealed considerable progress toward an understanding of plant–nematode interactions under natural conditions.

A current focus of the research community is to advance our understanding of phytonematode biology in sufficient detail to develop novel management methods. Progress in this aim is held back by a lack of functional genetic tools: forward genetics in the sedentary endoparasites is restricted to the root-knot nematode Meloidogyne hapla, and relies on natural variants as the source of mappable polymorphisms (Olaf Kranse et al, 2021) and reverse genetics is entirely reliant on RNA interference and is limited by the variable penetrance and stability of the effect. Despite these restrictions, meaningful progress recently has been made. There is nevertheless an expectation that the development of functional genetic tools would accelerate progress in understanding the biology of plant-parasitic nematodes, and thereby also indirectly the development of novel control solutions.

Nematoda, a diverse animal phylum that comprises an estimated million species, inhabit very broad ranges of ecological niches throughout earth. These animals, ranging from microscopic to a meter in size, are extremely successful in adapting different environments and have different lifestyles as free-living or parasitic to plants, animals and humans. As a result, nematodes have evolved to communicate with a wide variety of organisms that they live and interact with, including microbes, plants, insects, other animals, and nematodes of the same and different species. These communications play a key role in the mutualism, parasitism, predatory and prey, host and pathogen relationships between nematodes and other organisms and are critical to the ecological fitness of nematodes (Hsueh et al., 2014)

Pheromones are neuronal signaling molecules synthesized by various organisms and then excreted into the environment, where they typically stimulate individuals of the same species to react to environmental changes (e.g., temperature shifts, biological stimuli, or nutritional changes) (Jun Young Park et al., 2019). It is thought that most organisms, from prokaryotes to higher animals such as humans, can produce and use pheromones for communication between nonspecific individuals. In most cases, pheromones trigger neuronal events that are linked to various behavioral responses. The outcome of such neuronal stimulation includes the modulation of developmental and/or physiological programs that can support adaptation to new environments. 

Sex determination is one of the first aspects of nematode biology to be systematically characterized with genetic analysis and the depth of this analysis has made it a major topic in developmental biology. It is both fair and interesting to ask how general the Caenorhabditis elegans model is likely to be, both for other nematodes as well as for animals in general. As nematodes employ many different reproductive strategies, often related to parasitic life histories, one might expect similarly variable sex determination mechanisms. However, comparative studies of sex determination based on the C? elgans model have revealed both rapidly evolving and surprisingly well conserved features (Haag, 2005). This mixture of old and new (or slow and fast), along with its obvious relevance to evolution, ecology, and applied fields like agricultural and medical parasitology, has made the evolution of nematode sex determination an increasingly active research area.

7.1. Culturing of Nematodes

The need to use host plant tissue makes the culture of phytonematodes more difficult and expensive than the culture of fungi and bacteria. Relatively few species of phytonematodes have been cultured, compared to fungi and bacteria. Populations derived out of extraction procedures are often not sterile, an additional hindrance to many types of biological studies.

Axenic culture of free living, insect/animal parasitic nematodes in chemically defined or nondefined media has been achieved with some success, but development of similar culture system for plant parasitic nematodes has been less compared to former ones. However, these techniques form useful research tools in Nematology, which help procure clean nematodes, free of contamination for wide range of studies. Dual culture of nematode and plant tissue is aimed to establish plant parasitic nematodes in sterile cultures. Since phytophagous nematodes are obligate parasites they need a plant tissue as a food source. The term ’Gnotobiology’ is normally used to culture/study a single species of a nematode in the absence of other nematodes /organisms or in the presence of other known species. These techniques are generally considered as tedious and time consuming, require expertise and specialized equipments, nematodes are rarely found associated with only one or few other organisms and there is a need to get non-sterile soil and/or plant tissues, which does not go with gnotobiotic techniques.

Morphology based nematode taxonomy and biodiversity studies have historically challenged most biologists. In the past few decades, there have been efforts to integrate molecular methods and digital 3D image-capturing technology in nematode taxonomy, the former to enhance the accuracy of identification of such a taxonomically challenging group and the latter to communicate morphological data. While the employment of these two methods is growing in recent taxonomic, biodiversity and biogeographic studies, a movement to abandon traditional phenotypic identification methods altogether has emerged (Eyualem Abebe et al?, 2011). Proponents try to justify this trend by citing the challenging gap between the high estimated number of undescribed species and the limited ability of traditional taxonomy to accomplish the task of documenting such diversity.

Recent years have seen a rapid increase in the application of the various techniques of molecular, cytogenetical and serological approaches to problems in plant Nematology. The application of these approaches has been considered to problems of genetic variation and classical taxonomy. Two principal approaches are available. Firstly the use of “random” techniques which aim to sample the entire genome of the animaIs or populations of interest and which are mostly used in studies of genetic variation. Secondly “non-random “ techniques which target defined regions of the genome such as particular genes or gene products and are most frequently used in identification to species or sub-species level. The defined region ofthe genome may be the ribosomal genes, the mitochondrial genome or a conserved nuc1ear gene. Whilst analysis of these regions may allow information about taxonomic relationships to be gathered, focusing on one region of the genome imposes certain limitations on the experimenter.

Phytonematodes spend much of their lives inside or in contact with host tissue, and molecular interactions constantly occur and shape the outcome of parasitism.Eggs of these parasites generally hatch in the soil, and the juveniles must locate and infect an appropriate host before their stored energy is exhausted Components of host exudate are evaluated by the nematode and direct its migration to its infection site.Host plants recognize approaching nematodes before physical contact through molecules released by the nematodes and launch a defense response In turn, nematodes deploy numerous mechanisms to counteract plant defenses. A review by Shahid Siddique et al., (2022) focused on these early stages of the interaction between plants and nematodes and discussed how nematodes perceive and find suitable hosts, how plants perceive and mount a defense response against the approaching parasites, and how nematodes fight back against host defenses.

Future areas of emphasis for research and scholarship in nematode ecology are indicated by pressing agricultural and environmental issues, by new directions in applied nematology, and by current technological advances. Studies in nematode ecology must extend beyond observation, counting, and simple statistical analysis. Experimentation and the testing of hypotheses are needed for understanding the biological mechanisms of ecological systems. Opportunities for fruitful experimentation in nematode ecology are emerging at the ecosystem, community, population, and individual levels. Nematode ecologists will best promote their field of study by closely monitoring and participating in the advances, initiatives, developments, and directions in the larger f ield of ecology. Some major concepts in ecology have been explored including the dynamics of species interaction, the relationship between complexity and stability in ecosystems and the characterization of organisms as “r” or “K” selected (Howard Ferris, 1993). There has been some application of concepts of plant-herbivore coevolution and density-dependent regulation of populations.

Keeping the medical, ecological and economical importance of nematode phylum in mind, it is remarkable to see that nematode systematics is far from established. It has a long history of constant revision and there may be as many classifications as there are nematode taxonomists. Ferris and Ferris (1987) anticipated about the growing sense of excitement pervading systematics as new techniques make it possible a depth of understanding of phylogenetic relationships and affinities never before thought possible. They further stated that Darwin’s ‘genealogical taxonomy’ based on the concepts of descent with modification, is linked directly with two approaches to phylogentic inference, viz., phonetics and cladistics. In both these, patterns of descent take precedence over processes and in classifications based on these procedures, ‘grades’ and ‘gaps’ beloved by the evolutionary systematics are ignored and categories are usually of lesser importance. The phonetic approach deals with ‘natural classification’ based on overall similarity and the belief that the more characters a classification is based on, the more reliable it will be.

Nematodes are invertebrate roundworms that inhabit marine, freshwater and terrestrial environments. They comprise the phylum Nematoda or Nemata, which includes parasites of plants and of animals, including humans, as well as species that feed on bacteria, fungi, algae, and on other nematodes. Four out of every five multicellular animals on the planet are nematodes. All nematodes may look alike, the smallest being scaled down versions of the largest. The phrase tube-within-a-tube is a convenient way to think of nematode body structure and also a term used to refer to a major trend in the evolution of triploblastic metazoa. It refers to the development of a fluid-filled cavity between the outer body wall and the digestive tube. The nature of this body cavity has led to the grouping of metazoa into three grades, acoelomate, pseudocoelomate and eucoelomate. Nematodes together with Rotifera, Gastrotricha, Kinorhyncha, Nematomorpha, Acanthocephala and Entoprocta are traditionally grouped together as pseudocoelomates on the basis of possessing a body cavity that is not formed from the mesoderm or fully lined by peritoneum.

9.1 General

This chapter provides a comprehensive overview of the Life Cycle Assessment (LCA) of biochar production, focusing on the environmental impacts of its production and utilization. LCA is a systematic approach used to evaluate the environmental burdens and benefits of a product or process throughout its entire life cycle. Biochar, a carbon-rich material derived from the pyrolysis or gasification of biomass such as agricultural crop residue, forest waste, unusual wood waste, etc., has gained significant attention as a potential sustainable solid waste management, carbon sequestration, greenhouse gas mitigation, and soil amendment solution. However, understanding the environmental implications of biochar production is crucial for its sustainable implementation. The LCA approach is employed to evaluate biochar production’s different system boundaries and environmental performance, encompassing various stages such as biomass feedstock cultivation, the biochar production process, transportation, application, and end-of-life scenarios. This chapter explores the key elements of LCA, examines the methodologies employed in biochar LCA studies, and critically analyzes the environmental impacts associated with different biochar production pathways.

In order to assess the economic value of biochar, various factors need to be taken into account, including production expenses, market demand, potential income sources, and cost reductions. The economic worth of biochar can be determined by considering these factors.

8.1 Production costs

The cost of producing biochar can differ based on various factors such as the type of material used, the scale of production, the method of pyrolysis used, and the particular operational conditions.

i. Feedstock costs

• The cost of acquiring or producing biomass feedstock includes purchasing agricultural residues, forestry waste, or other biomass sources. Additional costs such as land, seeds, fertilizers, and harvesting should be considered using dedicated energy crops.

7.1 General

Science and technology have been advancing rapidly, but unfortunately, their impact has been largely negative. Pollution is not the only issue we face today, but also the depletion of fossil fuels, climate change, and excessive land degradation. The persistent presence of heavy metals in soil poses a significant threat to our planet. Many efforts have been made to remediate soils contaminated with toxic chemicals.

There are several issues that need urgent attention at the moment, and a solution is necessary to tackle them all at once. Biochar is a great option because of its numerous benefits, such as a large surface area, high porosity, high carbon sequestration rate, excess nutrients, and potential to produce syngas and bio oil. By using biochar, we can reduce the bioavailability of pollutants in soil, combat climate change by sequestering carbon, and reduce the use of fossil fuels by producing syngas and bio-oil. Additionally, biochar improves soil health, which can contribute to food security.

6.1 Characterization of solid pyrolysis products

 Biochar can be characterized for its utility as well as its energy usage. Thevarious characteristics of biochar can be categorized as follows:

i. Proximate analysis

 •            Raw moisture

 •            Hygroscopic moisture

 •            Volatile matter

 •            Ash content

 •            Fixed carbon content

 ii. Ultimate analysis

 •            Elemental compositions (C, H, N, O, S)

 iii. Physical properties

 •            True density

 •            Bulk density

 •            Porosity

 •            Dry matter

 •            Particle size

 •            Pore size

 •            Pore volume

 iv. Chemical properties

 •            pH-value

 •            Electrical conductivity

 •            Carbonate content

The classification of pyrolysis processes plays a crucial role in understanding and categorizing the various methods used for thermal decomposition of biomass, waste materials, and carbonaceous feedstocks. This chapter provides an in-depth overview of the classification schemes employed in the field of pyrolysis. It discusses the different factors considered for classification, including temperature, heating rate, feedstock, reactor configuration, and operating conditions. Understanding these classifications is essential for selecting the appropriate pyrolysis process based on specific objectives, feedstock availability, and desired product yields.

This chapter focuses on the design and performance evaluation of pyrolyzers, which are key components in producing biochar, biofuels, and other valuable products through the pyrolysis process. Pyrolyzer play a crucial role in converting biomass feedstock into desired end products, and their design greatly influences process efficiency and product quality. The chapter discusses various aspects of pyrolyzer design, including reactor types, operating conditions, heating methods, feedstock characteristics, and process optimization. It also explores performance evaluation methods to assess pyrolyzer efficiency, product yield, biochar quality, and energy consumption. The information presented in this chapter aims to provide researchers, engineers, and practitioners with valuable insights into pyrolyzer design and performance evaluation for efficient and sustainable biomass conversion.

3.1 Overview of pyrolysis technology

Pyrolysis is a process derived from the Ancient Greek words “pyro” meaning f ire and “lysis” meaning separation, which means separating by using heat or fire. This is a thermochemical conversion process that converts biomass into a useful intermediate liquid product, which can be further transformed into a hydrocarbon biofuel, oxygenated fuel additive, or substitute for petrochemicals. Pyrolysis is similar or overlapping with other thermochemical processes such as cracking, carbonization, torrefaction, destructive distillation, dry distillation, and thermolysis. It is not related to gasification, which involves chemical reactions with a gasification medium.

Biomass is a renewable and valuable resource that has many uses, including energy production, biofuels, and bioproducts. It’s made up of a complex mixture of organic compounds like cellulose, hemicellulose, lignin, proteins, fats, and minerals, which can vary depending on the source and type of biomass. Understanding the composition and properties of biomass is essential for efficient use. In this chapter, we’ll provide an overview of how to analyze and evaluate biomass as a renewable energy source. We’ll cover various techniques and methodologies for analyzing biomass, including physical, chemical, and thermal properties. We’ll discuss the importance of biomass composition, moisture content, ash content, elemental analysis, calorific value, and other relevant parameters when assessing its suitability for energy conversion. Additionally, we’ll highlight the significance of biomass characterization in understanding biomass feedstock variability and its impact on energy production efficiency, environmental performance, and sustainability.

1.1 General

Biomass is a renewable and plentiful energy source with enormous potential to meet the rising demand for clean and sustainable energy solutions. This chapter offers an overview of biomass and its conversion technology, which includes various techniques to transform biomass into useful forms of energy like heat, electricity, and biofuels.

Biomass is an organic substance obtained from plants, animals, and microorganisms. It can be utilized as a source of energy or as a basic material in various applications. The energy in biomass is derived from photosynthesis, during which plants convert sunlight, carbon dioxide, and water into organic matter. Since it comes from living or recently deceased organisms, it is a sustainable and renewable resource. In terms of carbon dioxide emissions, biomass is considered a carbon-neutral energy source because the carbon dioxide produced during combustion is offset by the carbon dioxide absorbed by plants during their growth.

Etiology

It is a pruritic transmissible dermatological condition caused by Cheyletiella yasguri, one of the surface living mite.

It is commonly spread by coitus and also by licking and rubbing of the affected area. It is more common in active breeding dogs.

It is more common in dogs and frequently seen in the skin and oral cavity. It usually occurs in 5 to 11 years old dogs. Benign melanomas are solitary, circumscribed, alopecic firm and freely movable blue black coloured nodules in the skin. Malignant melanoma is sessile, polypoid to plaque like grey or brown black coloured granules in the skin with ulceration.

It is more common in old dogs and also mostly observed in the cutaneous region of trunk and legs.

Grossly, the tumour appears as solitary to multiple masses as white, pink to red, smooth, and raised nodules. Alopecia and ulceration are also observed. It is mostly observed in the pinna and digits.

Cutaneous lymphoma has two types viz. Epitheliotropic and non- epitheliotropic. Epitheliotropic lymphoma (mycosis fungoides) is mostly seen in the skin which is characterized by erythema, pruritus, alopecia, scaling, erosion and ulceration. It mostly occurs in plaques or nodules (Figure 80). Non-epitheliotropic lymphomas are observed as nodules. They are usually sheets of nodules seen in the deep dermal and subcutaneous tissue.

Histiocytoma is more common in younger dogs below two years of age. It is also observed in all age groups.

Gross examination

Grossly, histiocytoma appears as light pink, smooth to firm, raised round (Figure 77) masses. Alopecia and ulceration are also observed.

The mast cell tumors are focal or multicentric in nature in the skin and occasionally in the internal organs. It is more common in middle aged dogs without sex predilection.

Hemangiopericytoma is a tumor originating from pericytes. It is mostly seen in distal extremities due to elevation of blood pressure in that area. It is mostly seen in skin and subcutaneous tissue around the joint region.

Haemangiosarcoma is a malignant tumour of vascular endothelial cells. It is more commonly seen in aged dogs.

It is a benign tumor of endothelial cells. It is a common tumor of dogs. Solar induction is one of the cause for development of this tumor.

Liposarcoma is a rare malignant tumor of adipocytes in dogs. Grossly, the neoplastic cells are arranged in less circumscribed to large firm to hard (Figure 67) nodules on the dermis and the nodules noticed on the subcutis are grey white in color.

Lipoma is a benign tumor of adipocytes and it is more common in dogs and mostly observed in the trunk, thigh and proximal limbs with rare presence of ulceration.

The tumour is most commonly seen in adult and aged dogs. It occurs anywhere on the body surface. It is due to trauma, chronic inflammation, vaccination, genetic predisposition etc.

Benign/malignant tumor may arise from fibrous tissue. Fibroma mostly occurs in the dermis or subcutis of limbs and head.

Perianal gland tumors originate around the anus of the dog, upper and lower part of the tail and prepucial area. Retention of gonadal hormone receptors is one of the causes for perianal gland adenoma and adenocarcinoma.Cytology reveals the presence of hepatoid cells with centrally placed nuclei and prominent nucleoli. A small flattened reserve cells or resting cells are also noticed around the hepatoid cells. In malignant variety, the cells containing large vesicular nuclei, eosinophilic granular cytoplasm and binucleated cells, mitotic figures, reserve cells, moderate stroma and lymphoplasmacytic infiltration are seen (Figure 55 & 56).

The tumor arising out of ceruminous gland might develop subsequent to chronic otitis externa in dogs. It may be a benign or malignant tumor of ceruminous gland epithelium. It is more common in the external ear canal of dogs.

It is one of the common epithelial skin tumors. It is more common in head but also recorded in other sites such as back, tail, limb, trunk and eyelids. It is a tumor of sebaceous cells. In sebaceous gland adenoma, it has predominantly sebocytes with less of basaloid reserve cells while in sebaceous epithelioma variant, more of basaloid reserve cells are seen with less of sebocytes.

This is a benign/malignant tumor originating from the apocrine and eccrine sweat glands in dogs. It is usually recorded in the head and neck region but it may occur on any site.

Cytology reveals clusters of moderate to severe exfoliated neoplastic cells which contain round nuclei slightly eccentric in position in the light pale granular cytoplasm. In carcinomas, clusters of basophilic epithelial cells had large nuclei and prominent nucleoli. It may also contain one or two large secretory droplets.

Basal cell carcinoma is a locally malignant epithelial tumor of skin which is mainly composed of basal cells. This may occur due to mutation in the tumor suppressor gene p53 and chronic exposure to UV radiation.

Cytologically, neoplastic cylindrical columnar cells, are moderate to high in number. The neoplastic cells are small to medium sized with round to spherical nuclei with basophilic cytoplasm (Figure 42 & 43).

This tumour may occur due to specific mutation in the p53 tumor suppressor gene. This is one of the most common epithelial tumor of dog. It is classified into well differentiated, moderately differentiated and poorly differentiated tumour.