Intellectual property rights (IPR) are designed to allow novel technologies to be available so that the scientist or company receives a reward for the initiative established. Intellectual property possessions can be any codified knowledge, innovation, or anything of actual or potential economic value that has arisen from rudimentary research, analysis, and manipulation of biological systems, industrial application, or for commercial use.
The various types of biotechnological inventions may be grouped into the following-
Approaches/processes of generating useful products.
Numerous products, for example- Antibiotics, vitamins, etc.
Applications of various processes/products, for example, the use of a promoter sequence to regulate gene action.
DNA sequences and the proteins.
Strains of microorganisms, cell lines are obtained by genetic modification.
Methods for genetic modification of organisms.
Patenting of Genes and DNA Sequence–
An artificially synthesized gene is patentable in almost all countries. A patented gene holds exclusive rights to the specific DNA sequence. Once patented, the holder of the patent dictates how the gene needs to be used (whether commercially or clinically) for a minimum of 20 years from the date of the patent. In the USA, genes isolated from the organisms are patentable; gene aroA (shows glyphosate resistance) isolated from a mutant bacterium was the first to be patented. For a patent to be granted in India, it should not be covered in the negative list in Section 39 which provides an extensive list of what are not the inventions under the Indian Patents Act. The act came into force in 1972 amending the Patent act,1970.
The three conditions in order to fulfil the rules imposed by Indian patent act are :
• It should be a novel creation
• It should involve an inventive step for the mankind
• There should be various industrial applications.
Ananda Mohan Chakrabarty got the first US patent for a genetically modified organism in 1981. He discovered a method for cross-linking in such a way that it fixed all the 4 plasmids to a much stabler microbe called Pseudomonas putida capable of consuming 2-3 times faster than previous strains. Its unique characteristic was hydrocarbon degradation, therefore the name was given as “multiplasmid hydrocarbon-degrading Pseudomonas”/superbug. Prof. Chakrabarty’s momentous research has since paved the way for many patents on genetically modified micro-organisms and other life forms for the coming years.
Can Life forms be patented?
The main arguments in favouring the patent of genetically modified life forms are
-They perform novel and useful functions
-They generate economic benefits
-Their production requires large financial and technical innovative inputs
However, the chief objections are usually based on ethical, moral, and religious considerations such as
-They are products of nature and hence should not be fiddled with
-Their genetic modification does not prove an industrial invention
-The inventions cause cruelty to animals
But in 1985, a patent was granted in the USA for a maize plant by overproducing tryptophan through plant tissue culture. Later, in 1988, a genetically engineered mouse called “OncoMouse” was the first mammal to be patented. It was primarily used for cancer research. The animal designed by Philip Leder and Timothy A Stewart of Harvard University used to carry a specific gene called known as an activated oncogene. The activated oncogene increased the mouse’s resistance to cancer, and thus the mouse is a promising model for cancer research. The patenting of OncoMouse, and the extensiveness of the claims made in those patents, were well-thought-out to be unreasonable by many of their colleagues. But, amid the controversies, it was finally patented in 1992.
Prions are a misfolded pathogenic form of proteins that have the ability to transfer its misfolded form to the normal shape of the same protein thereby, folding abnormally. The word “prions” was coined in 1982 by Stanley B. Prusiner and it is a derivative name from “proteinaceous infectious particle”. They are responsible for transmitting several lethal neurodegenerative diseases, particularly in human and few other animals. It is believed that the abnormal 3D configuration of the misfolded protein contains infectious properties resulting in turning the surrounding normal proteins into abnormal ones. These infectious agents are very different from other known infectious agent such as virus, bacteria, fungi as, all of them contain a genetic material (either DNA or RNA).
Prions are made up of proteins called as PrP that is generally found throughout the body of healthy humans and animals. But, PrP found in of the infectious prion has a different structure and is very resistant to proteases. Proteases are those enzymes that usually breakdown the proteins in the body. The healthy PrP is known as PrPc while the infectious one is known as PrPSC meaning ‘scrapie’, a disease of prion occurring in sheep. The structure of PrPc is well defined while the structure of PrPSC is poorly defined.
There were two hypotheses explaining the replication of prions. First one was a heterodimer model in which an assumption was made that a single PrPSC molecule binds to PrPc and further catalyzes its conversion to PrPSC. These PrPSC can go and further convert PrPc. But it was difficult to explain the spontaneity of the two molecules, hence it was disproved. Second model assumes that PrPSC exists as fibrils and its ends bind to the PrPc and further convert it into PrPSC. But an exponential growth was observed along with quantity of infectious particles thereby, explaining a breakage in the fibril.
THEIR ROLE IN NEUROGENERATIVE DISEASE–
Prions cause neurogenerative disease by aggregating from outside inside the nervous system. This forms plaque known as amyloids that completely changes the tissue structures. This is visualized by “holes” in the tissue resulting in a spongy-like structure in the neurons. The incubation period of prion-related disease is (5-20 years), but once symptoms appear, they progress very quickly. The result might be brain damage or death. The structure of infectious prions usually is similar to every species and hence there is a little chance that it will get transmitted. However, the human prion disease Creutzfeldt- Jakob is said to be the prion that usually infects cattle, knowans as mad cow disease which severely infects the cattle and is transmitted through contaminated meat. Also, these proteins have been implicated in the ontogeny of neurodegenerative diseases such as Alzheimer’s disease, Parkinson’s disease, Hunington disease, and frontotemporal lobar degeneration with ubiquitin-positive inclusions (FTLD-U).
All prion-related diseases do not have a cure and are fatal. The clinical trials for the same have not produced successful results yet because of the rarity of disease. Although, some experiments built genetically engineered cattle and mice which lacked the gene which is necessary prion production, therefore, building research on the same.
The Spemann-Mangold organizer are a consortium of cells that are required for the commencement of the neural tissue during the development of an amphibian embryo. Hilde Mangold, the then doctorate student along her mentor Hans Spemann, first published this work in 1924. This discovery gave so much scope to the developmental biology field, that this became one of the few doctoral theses to have won the Nobel prize in 1935. They showed that, of all the tissues in the early gastrula stage, only one has its fate determined. The overview of this organizer proved that destiny of the cells can be altered and influenced by factors from other cell clusters.
This self-differentiating tissue is the dorsal lip (the dorsal bordering region of the blastopore, which acts as the centre of differentiation) of the blastopore, the tissue derived from the grey crescent cytoplasm. When this dorsal lip tissue was transplanted into the so-called belly skin region of another gastrula, it not only continued to be blastopore tip, but also started the process of gastrulation and embryogenesis in the nearby tissues. Later on, two conjoined twins were formed instead of one.
In this experiment, Spemann and Mangold used two different pigmented embryos from two newt species- that is, the darkly pigmented Triturus taeniatus and the non-pigmented Triturus cristatus. Two different species were taken for this experiment for transplantation by Spemann and Mangold as it would be easier to identify which one was the host and donor tissues respectively. At first, the dorsal lip of an early T. taeniatus gastrula was removed and then implanted into the area of an early T. cristalus gastrula was supposed to turn into ventral epidermis. As predicted, the dorsal lip tissue invaginated (cleaved) showing the qualities of self-determination and disappeared under the vegetal cells. The egg is divided into two regions- the animal pole (top part of the egg) and the vegetal pole (bottom part of the cell). Usually, the genetic material and proteins are unevenly distributed among these two poles.
The donor tissue (the pigmented species) of newt then continued to self-differentiate and divide into a structure called chordamesoderm (notochord) and other mesodermal structures respectively which usually comes from the original dorsal lip.
Now, these newly made donor- derived mesodermal cells move forward for further participation in differentiation. As they are in movement, the host cells participate in the formation of new embryo. It creates organs that normally never would have formed before. In the secondary embryo, a somite could be seen containing both pigmented (that is, the donor) and the unpigmented (which is the host) tissues. What was more shocking was, the dorsal lip was able to interact strongly with the host tissue to form a fully formed neural tube derived solely from the host’s ectoderm. Back then, Spemann referred to the dorsal lip cells and their derivatives as the organizer. The reason being was because-
They could induce the host’s ventral tissue to change their fates to form a neural tube and a dorsal mesodermal tissue (most commonly the somite).
They could systematically organize host and donor tissues into a secondary embryo with clear anterior-posterior and dorsal-ventral regions.
Because, there are numerous inductions during the embryonic developments, this key induction wherein the progeny of dorsal lip cells induces the dorsal axis and the neural tube is traditionally called the Primary embryonic organizer.
Throughout the course of modern human history, the source some viral infections such as smallpox, polio, and the Spanish flu have been quite unknown to humans. They have been diseases which have had deadly effects on humanity. All that was known about these diseases was that they spread via person-to-person contact.
In the second half of the1800’s, Louis Pasteur postulated that the disease rabies was caused by a “living thing” which is in all probability, smaller than bacteria itself. This postulation of his led him to develop the first vaccine against rabies in 1884.
The initial discovery of the light microscope held some promise with respect to the observation of the agents causing such sever diseases however, it was found that the microscope could only be used to observe bacteria, protozoa and fungi. The size of virus was smaller than these agents and therefore, could not be observed under a microscope.
In the 1890’s, D. Ivanovski and M. Beijerinck showed that a disease caused in plants was cause by the tobacco mosaic virus which served as the first properly substantial revelation related to viruses.
This discovery was then followed by two other scientists, Friedrich Loeffler and Paul Frosch who isolated the virus that caused foot and mouth disease in cattle.
POSITION OF VIRUSES IN THE BIOLOGICAL SPECTRUM:
Viruses are extremely unique entities which are capable of infecting almost every single type of cell known including bacteria, algae, fungi, plants, animals and even protozoa.
Questions regarding the nature of viruses like their origination, their state of existence (alive or non-living), their distinct biological characteristics etc. are still very dominant in the scientific world.
Some ideas that have been addressed are:
Viruses are considered to be the most abundant microbes on earth, in terms of number.
Viruses are considered to be obligate intracellular parasites that are incapable of dividing or multiplying unless they invade a specific host cell. They multiply by taking over the hosts genetic and metabolic machinery of the cells of the host.
They are said to have been in existence for billions of years and have arisen from the loose strands of genetic materials released by the cells. (This is one widely accepted theory. However, it too has faced some criticism.)
PROPERTIES OF VIRUSES:
Viruses are obligate parasites of protozoa, bacteria, fungi, animals, plants, and algae.
They have an ultramicroscopic size range from 20nm up to 450 nm in diameter.
They have a very compact and economical structure. They are not cellular in nature.
They are inactive macromolecules outside the host cell and only get activated inside the host cells.
Viral nucleic acid can be DNA or RNA but never both together.
Their basic structure consists of a protein coat (the capsid) surrounding the nucleic acid core.
They lack the enzyme and machinery for basic metabolic processes and synthesis of proteins.
Virus multiply by taking over the host machinery and genetic material.
THE GENERAL STRUCTURE OF VIRUSES:
Viruses represent the smallest infectious agents in the biological world. (with a few exceptions)
They lie within the ultramicroscopic size range with sizes usually less than 0.2 micrometre. They are so small that one requires an electron microscope to detect or examine their structure.
The animal size range may vary from parvoviruses (which are around 20 nm in diameter) to megaviruses and pandoraviruses (which are up to 1000nm in width) that may be as big as small bacteria.
Some viruses which are cylindrical may be relatively long (around 800nm) but have a very narrow diameter (around 15nm).
Negative staining using an opaque salt in combination with electron microscopy can be used for the observational studies of viruses.
STRUCTURAL COMPONENTS OF A VIRUS:
Viruses have a crystalline appearance due the occurrence of regular, repeating molecules. A number of purified viruses even form large aggregates and crystals when subjected to special treatments.
The general plan of all viruses or the general architecture is quite simple. Almost all viruses contain a protein coat or the capsid which encloses the viral genome which may be a DNA or an RNA sequence. Apart from this, viruses only contain those parts which are needed to invade and take control over the hosts cellular machinery.
Some important terms are:
Capsid: The outer covering or the shell of the virusthat surround the central nucleic acid core of the virus.
Nucleocapsid: The outer shell along with the nucleic acid core is called the nucleocapsid.
Naked viruses: Viruses that do not contain a nucleocapsid layer are called naked viruses.
Enveloped viruses: Some virus classes possess an additional covering which is external to the capsid which is called an envelope. This envelope structure is usually a piece of the hosts cell membrane. These types of viruses are called enveloped viruses.
THE VIRAL CAPSID:
The capsid layer of the virus, when magnified immensely, shows the appearance of small, prominent, geographic structures. These structural subunits of the capsid are called capsomeres.
These capsomeres are capable of self-assembling into the finished capsid structure. Depending on the shape and assembly of the capsomeres, the resulting structure can be of two types;
Helical: Helical capsids have rod shaped capsomeres that bind together to form a structure similar to hollow discs (like a bracelet). During the process of formation, these discs link together to form a continuous helix.
Icosahedral capsids: An icosahedron is a three-dimensional, 20-sided figure with 12 evenly spaced corners. Some viruses also show such an arrangement in this shape.
Complex viruses: Such viruses may have a specific head, a neck and other structures specific for the invasion of host cells. The most common examples of such viruses is phage viruses.
THE VIRAL ENVELOPE:
Enveloped viruses, when released from the host cells sometimes carry forward a piece of the hosts cell membrane with them in the form of an envelope. Although it is derived from the host, the envelope is different in the virus because the normal proteins of the host get replaced with the viral proteins.
At the center of the viral structure, within the capsid lies the viral genome which may be single or double stranded. The genetic material may be RNA or it may be DNA but it is never both even in viruses. The genome may be a few hundred to thousands of base pairs long.
The potential for spreading through drinking water is the emerging pathogenic bacteria of concern outlined here, but they do not correlate with the existence of E. Coli or with other measures of the consistency of drinking water widely used such as coliform bacteria. There are no satisfactory microbiological markers of their existence in most cases. To understand the real nature and dimension of the diseases caused by water polluted with these bacteria and the ecology of these pathogens, further studies are required.
Mycobacterium Avium Complex (Mac):
The complex Mycobacterium avium (Mac) consists of 28 serovars of two species: Mycobacterium avium and Mycobacterium intracellular. With the discovery of disseminated infection in immunocompromised individuals, especially people with HIV and AIDS, the Mac species’ significance was recognized. MAC members are deemed to be opportunistic human pathogens. A wide range of environmental sources, including coastal waters, rivers, lakes, streams, wetlands, springs, soil, piped water supplies, plants, and house dust, have defined Mac species. Mac species have been isolated from the delivery systems of natural water and drinking water in the USA. The ubiquitous existence of Mac organisms stems from their ability under varied conditions to thrive and evolve. Mac species can proliferate at temperatures up to 51°C in water and expand over a broad pH range in natural waters. These mycobacteria are incredibly resistant to the use of chlorine and other chemical disinfectants in drinking water care. Standard drinking-water treatments may not remove Mac species but may substantially reduce the numbers present in the source water to a level that poses a negligible risk to the general public if it is running satisfactorily. In delivery systems, the entryway for these mycobacteria is through leaks. For their continued presence in distribution systems, the growth of Mac organisms in biofilms is probably significant.
Slow-growing mycobacteria can be present in the surface biofilm at densities higher than 4,000 per cm2, producing a potentially high exposure level. The signs of Mac infections result from either respiratory or gastrointestinal colonization, potentially spreading to other places in the body. Exposure to Mac species may occur through the consumption of contaminated foodstuffs, the inhalation of air containing contaminated soil particles, or through touch or ingestion, aspiration or aerosolization of the organisms containing drinking water. Unlike gastrointestinal pathogens, where E. No appropriate indicators have been identified to signal increasing Mac species concentrations in water systems, and coli can suggest possible presence.
As a significant etiologic agent for gastritis, Helicobacter pylori has been cited and has been involved in the pathogenesis of duodenal ulcer and peptic disease and gastric carcinoma. Most people who are infected by this pathogen, however, remain asymptomatic. Using methods based on history, H. There has been no isolation of pylori from environmental sources, including water. Molecular methods have, on the other hand, been useful in detecting the pathogen.
Fluorescence in situ hybridization (FISH) has been successfully used to detect this pathogen in drinking water delivery systems and other water bodies. To detect the presence of H, a polymerase chain reaction was also used. Pylori DNA in drinking water, especially biofilm-associated. In biofilms for drinking-water, H. Pylori cells lose culturability rapidly and enter a viable but non-culturable state. Cells persist for more than one month in these biofilms, with densities exceeding 106 cells per square cm. It remains unclear how the organism is transmitted. Nevertheless, the fact that it has been oral-oral or fecal-oral transmission is demonstrated by recuperation from saliva, dental plaques, stomach, and fecal samples. Water and food tend to be of less immediate significance, but they can still play a significant role in improper sanitation and hygiene.
Over the past years, A. Hydrophila has received attention as an opportunistic pathogen for public health. The elderly, children under the age of five, and immunosuppressed persons may play a significant role in intestinal disorders. Gram-negative, non-spore-forming, rod-shaped, facultative anaerobic bacilli belonging to the Aeromonadaceae family are Aeromonas hydrophila. Even though the dominant species is typically hydrophila, whereas other aeromonads, such as A.Sobria, and A.Caviae were isolated from human feces and water sources. Species of Aeromonas, including A. Hydrophila, in the field, are ubiquitous. It is also segregated from food, potable water, and aquatic ecosystems. Concentrations of Aeromonas spp. in safe rivers and lakes Typically, 102 colony-forming units (CFU)/mL are around. In general, groundwater contains less than 1 CFU/mL. It was noticed that drinking water immediately leaving the treatment plant contained between 0 and 102 CFU/mL. Drinking water can show higher concentrations of Aeromonas in delivery systems due to the growth of biofilms. With Aeromonas spp. growth was observed between 5° – 45° C.
A. Hydrophila is immune to standard treatments with chlorine and is likely to live within biofilms. Ingestion of infected water or food or touch of the organism with a break in the skin are the typical routes of infection suggested for Aeromonas. A potential source of pollution for human beings may be drinking or natural mineral water. There was no recorded person-to-person transmission.
For life, water is essential. There must be sufficient, stable, and usable supplies available to everyone. Improving access to clean drinking water will lead to substantial health benefits. Most individuals fail to gain access to clean water. The supply of clean and filtered water to each house may be the standard in Europe and North America, but access to clean water and sanitation is not the rule in developing countries, and water-borne infections are common. There is no access to better sanitation for two and a half billion people, and more than 1.5 million children die from diarrheal diseases each year. According to the WHO, the mortality of water-related illnesses exceeds 5 million individuals per year; more than 50 percent of these are microbial intestinal diseases, with cholera standing out first and foremost.
A prominent public health concern in developing countries is acute microbial diarrheal diseases. Those with the lowest financial resources and the worst hygiene services are those affected by diarrheal diseases. Children under five, especially in Asian and African countries, are the most affected by water-borne microbial diseases. Often affecting developing countries are microbial water-borne diseases—the most severe water-borne bacterial illnesses.
Table 1: Some bacterial diseases transmitted through drinking water.
1. Microorganisms such as viruses and bacteria in water bodies, which are used as a proxy to determine the existence of pathogens in that area, are indicator organisms.
2. It is preferred that these microorganisms are non-pathogenic, have no or limited water growth, and are consistently detectable at low concentrations.
3. In larger populations than the related pathogen, the indicator species should be present and preferably have comparable survival rates instead of the pathogen.
4. In the monitoring of water quality, different indicator species can be used, and the efficacy of predicting pathogens depends on their detection limit, their tolerance to environmental stresses and other pollutants.
Tiny, curved-shaped Gram-negative rods with a single polar flagellum are Vibrio. Vibrios are optional anaerobes capable of metabolism that are both fermentative and respiratory. For all animals, sodium promotes the development and is an absolute prerequisite for most.
1. The majority of plants are oxidase-positive, and nitrate is reduced to nitrite.
2. Pili cells of some microbes, such as V. cholerae, V. parahaemolyticus, and V. vulnificus, have structures consisting of TcpA protein.
3. The production of TcpA is co-regulated with cholera toxins expression and is a primary determinant of in vivo colonization.
4. Several species of Vibrio can infect humans. The most significant of these species is, by far, V. cholerae.
5. Several forms of soft tissue infections have been isolated from V. alginolyticus.
6. The cells of Vibrio cholerae will expand at 40°C at pH 9-10.
7. The presence of sodium chloride stimulates growth. Vibrio cholerae is a bacterial genus that is very diverse.
8. It is split into 200 serovarieties, distinguished by the lipopolysaccharide (LPS) structure (O antigens). Only O1 and O139 serovarieties are involved in true cholera.
9. Gastroenteritis may be caused by many other serovarieties, but not cholera.
10. Biochemical and virological features are the basis for the differentiation between Classical and El Tor biotypes.
1. The incubation period for cholera is 1-3 days.
2. Acute and severe diarrhea, which can reach one liter per hour, characterises the disease.
3. Patients with cholera feel thirsty, have muscle pain and general fatigue, and display anuria symptoms accompanied by oliguria, hypovolemia, and hemoconcentration.
4. In the blood, potassium decreases to deficient levels. With cyanosis, there is circulatory collapse and dehydration.
Several factors depend on the seriousness of the illness:
(a) personal immunity: both previous infections and vaccines can confer this immunity;
(b) inoculum: disease arises only after the absorption of a minimum quantity of cells, approx. 108.
(c) Gastric barrier: V. cholera cells like simple media and the stomach is also an adverse medium for bacterial survival, usually very acidic. Patients that take anti-acid drugs are more vulnerable than healthy patients to infection.
(d) Blood group: persons with O-group blood are more vulnerable than others for still unexplained causes.
5. In the absence of treatment, the cholera-patient mortality rate is approx—fifty percent.
6. The lost water and the lost salts, mostly potassium, must be replaced.
7. Water and salts can be administered orally during light dehydration, but rapid and intravenous administration is mandatory under extreme conditions.
8. Presently, doxycycline is the most effective antibiotic. In some instances, if no antibiotic is available for treatment, the administration of salt and sugar water will save the patient and help with recovery.
9. Two significant determinants of infection exist:
(a) the adhesion of bacterial cells to the mucous membrane of the intestine. It depends on the presence on the cell surface of pili and adhesins;
(b) development of a toxin from cholera.
1. Gram-negative motile straight rods include the genus Salmonella, a member of the family Enterobacteriaceae.
2. Cells are oxidase-negative and positive for catalase, contain D-glucose gas, and use citrate as a sole source of carbon. There are many endotoxins in Salmonellae: O, H, and Vi antigens.
3. S. Subsp enterica. enterica serovar Enteritidis is the most widely isolated serovariety worldwide from humans. Other serovarieties can, however, be prevalent locally.
4. A fermented juice historically extracted from the palm-tree was the source of insulation.
1. Two forms of salmonellosis can be pathogenic to humans:
(a) typhoid and paratyphoid fever (not to be confused with rickettsia-induced typhus disease);
2. Low infection doses are sufficient to cause clinical symptoms (less than 1,000 cells).
3. There are different clinical signs of salmonellosis in newborns and children, from a severe typhoid-like disease with septicemia of a range to a mild or asymptomatic infection.
4. The infection is commonly spread through the hands of staff in pediatric wards.
5. Ubiquitous Salmonella serovars, such as Typhimurium, are often caused by food-borne Salmonella gastroenteritis.
6. Symptoms such as diarrhea, vomiting, and fever occur about 12 hours after consuming infected food and lasts 2 to 5 days.
7. Spontaneous healing typically happens. All kinds of food can be associated with Salmonella.
8. The prevention of food-borne Salmonella infection is focused on the prevention of contamination, the prevention of food-borne Salmonella multiplication (persistent storage of food at 4oC), and where possible, the use of pasteurization (milk) or sterilization (other foods).
9. When infected with fertilizers of the fecal origin or washed with polluted water, vegetables and fruits can carry Salmonella.
10. The incidence of typhoid fever decreases as a country’s development level grows, such as pasteurization of milk, dairy products, and controlled water sewage systems.
11. The risk of fecal contamination of water and food remains high where these hygienic conditions are absent, and so is the occurrence of typhoid fever.
Bacillary Dysentery or Shigellosis:
Shigella are members of the Enterobacteriaceae family that are Gram-negative, non-spore-forming, non-motile, straight-rod-like. Without gas production, cells ferment sugars. There is no fermentation of salicin, adonitol, and myo-inositol. Cells do not use citrate, malonate, and acetate as the primary source of carbon and do not create H2S. It is not decarboxylated with lysine. Cells are oxidase-negative and positive for catalase. Members of the genus Shigella have a complex antigenic sequence, and their somatic O antigens are the basis of taxonomy.
1. The incubation time for shigellosis is 1-4 days.
2. Typically, the illness starts with fever, anorexia, tiredness, and malaise. Patients exhibit irregular, low-volume, sometimes grossly purulent, bloody stools, and abdominal cramps.
3. Diarrhea progresses to dysentery after 12 to 36 hours, with blood, mucus, and pus appearing in feces that decrease in volume (no more than 30 mL of fluid per kg per day).
4. Even though the molecular basis of shigellosis is involved, the colonic mucosa’s penetration is the initial phase in pathogenesis.
5. Degeneration of the epithelium and acute inflammatory colitis in the lamina propria define Shigella infection’s resulting concentration.
6. Desquamation and ulceration of the mucosa eventually contribute to leakage into the intestinal lumen of blood, infectious elements, and mucus.
7. The colon’s water absorption is hindered under some circumstances, and the amount of stool depends on the flow of ileocecal blood.
8. As a consequence, normal, scanty, dysenteric stools can move through the patient.
9. The bacterium must first adhere to its target cell in order for Shigella to penetrate an epithelial cell.
10. The bacterium is usually internalized into an endosome, which is then lysed to obtain entry to the cytoplasm where replication occurs.
The study of living microbes that are suspended in the air is known as Aero microbiology. Such microbes are known as bioaerosols. There are significantly fewer microorganisms in the atmosphere than in the oceans and in the soil; there are still many microorganisms that can impact the atmosphere. With the help of wind and precipitation, these microbes have a chance to migrate long distances and increase the rate of infectious diseases caused by these microbes. In humans, animals, and plants, these aerosols are ecologically important because they can be associated with the disease. Microbes can suspend themselves in the atmosphere, where they can communicate and precipitate with the clouds and create specific shifts in the clouds.
The air has two microbial ecosystems.
1. High light intensities, extreme temperature fluctuations, low amount of organic matter, and a lack of water availability; characterize the atmosphere as a habitat, making it a non-hospitable environment for microorganisms and a generally inadequate habitat development.
2. In the lower regions of the atmosphere, however, large numbers of microbes are contained.
1. In the atmosphere, an apparent mass of concentrated watery vapor floating, usually well above the general ground level.
2. Clouds, with a pH ranging from 3 to 7, are also an acidic environment.
Sources of airborne microorganisms:
1. Air is not a favourable microbial growth environment because it does not provide adequate moisture and nutrients to sustain growth and reproduction, and there is also no indigenous flora growth in the air.
2. Quite a range of sources responsible for introducing microbes into the air have been identified and researched.
3. The most popular of these is dirt. Microbes are suspended in the air with wind flow and remain there and often accumulate.
4. Microbes are often released into the air by human activities such as digging, sloughing, and running.
5. Microbes are often released into the air through air currents and splashes of water.
6. Besides, air currents strip plant and animal pathogens from their surfaces and disperse them across the atmosphere.
7. In contrast to animal pathogens, plant pathogens can spread more quickly. For example, a gamine flies over a thousand kilometers with Puccini spores.
Examples of airborne plant pathogens:
Examples of airborne animal pathogens:
8. Human beings are the primary cause of the introduction of bacteria into the air.
Examples of airborne human pathogens:
9. The most comprehensive source is human activity. The pathogenic bacteria in the human respiratory tract and the mouth’s microbes are continuously released into the air, but they cough, sneeze, and laugh.
Depending on the size and moisture content, the microbes released into the air come in three forms. Those are the three forms:
2. Nuclei Droplets
3. Dust that is contagious
Droplets: As we sneeze, millions of droplets are released, and mucus is expelled from about 200 miles away. Such droplets are water droplets that hold microorganisms if a diseased individual releases them. Saliva and mucus comprise these droplets. Most of the microorganisms they transport are from the respiratory tract. The droplet size determines how long microorganisms live on the droplet. Large-sized droplets settle quickly in the air. The source of these droplets carrying the microorganism can be a source of infectious disease.
Water particles emitted 1 to 5 micrograms in diameter during sneezing and coughing. For respiratory disorders, droplet nuclei are known to be the raw material. On its surface, it contains saliva and mucus. They are stuck in the air for a more extended period because of their small scale. Droplet nuclei, if the bacteria are the constant source of bacterial infections, are known to be
The present remains viable on its surface. The viability of bacteria depends on physical conditions, such as humidity, sunlight, moisture, and droplet size.
By bed making, holding a handkerchief, working with a patient with dried secretion, digging and ploughing, these dust particles are released into the air. Microorganisms adhere to these droplets’ surface and are then suspended by the above techniques to dry them. There is a more significant size of dust particles laden with bacteria and settle down in the air. Two forms of droplets cause airborne diseases.
a. Droplet infection due to droplets with a diameter greater than 100-micron meters.
b. Any dried droplet residues cause airborne infections.
Infection with droplets remains localized and concentrated, while airborne infection may be long-distance. Microorganism can grow on dust particles for a more extended period. It is proven harmful in hospitals and laboratories when closed bottles of dried specimens are opened, and cotton plugs are removed from the bottles.
Factors influencing airborne microbes:
Factors that influence microbial survival in the atmosphere are
c. Content of Nutrients
d. pH and Acidity
1. The main factor in regulating the growth of microbes in the air is temperature.
2. High temperatures hinder the production of microbes and often denature the microbes’ structural conformation.
3. Very few microbes can live and withstand high temperatures, i.e., extremophiles.
4. Likewise, when ice crystal formation occurs, shallow temperatures are also not ideal for microbial growth.
5. Humidity has a role in preserving the development of airborne microbes.
6. Gram-Bacteria associated with aerosols tend to live in low humidity for a more extended period.
7. The abundance of nutrients in the atmosphere is lower, so it does not help microbial growth.
Microbial growth is defined as the increase in cell number (rather than cell size) by asexual reproduction process called, binary fission. Cell division results in the growth of the cells in the representative population. Bacteria and archea undergo asexual reproduction, while fungi and higher organisms also exhibit sexual reproduction and increase in cell size is also considered as growth in higher organisms.
Binary fission is the process of asexual reproduction method followed by most bacteria. In this process, initially a single cell starts doubling its machinery, including genetic material, enzymes and other essential components. Then a wall like structure, called septum forms in middle, dividing the cell into two equal halves, each half receives a set of genetic material and other essential components needed for life. A typical bacterium like Escherichia coli takes around 20 minutes to divide into two and is called a cell cycle. Multiple fission, budding and sporulation are some less frequent kinds of cell division shown by bacteria. The bacterial cell division by binary fission was shown in figure 1.
Bacterial growth curve
. When conditions favor (nutrients, environment, temperature etc), bacterial growth rate is rapid. When the bacteria undergo any change in their niche environment, growth rate is effected that might lead to dormancy or death. E.coli is taken as a model organism as it has very small life cycle of 20 minutes and easy to grow in the lab for variety of biological studies
E.coli can be grown in a glass test tube and its growth phase can be studied in a predictable approach. A typical bacterium under controlled conditions will exhibit four phases as follows:
i) Lag phase
ii) Log phase
iii) Stationary phase
iv) Decline/death phase
Lag phase: Bacteria in this phase will be in adaptive mode, trying to adjust to the surrounding new environment. If cells are in damaged state or transferred to entirely new environment, bacteria takes long time to adjust and the lag phase becomes lengthy. If the bacteria are exposed to familiar kind of environmental conditions with suitable nutrients, temperature etc, growth starts rapidly. In this case, lag phase takes less time.
In lag phase, cells will rejuvenate themselves; synthesize enzymes, metabolites, RNA etc that are required for the growth. Cells try to repair if there is any damage that was there in the cells.
Log phase: This phase is also called exponential phase. Cells will be in active mode and increase in number by multiplication, where 1 cell becomes 2, 2 becomes 4 etc. Rapid growth of the cells will result under ideal conditions, while showing slower growth is very rare depending upon the nutrient or other conditions. Cells in this phase are the healthiest and in the most active form. Exponential stage cells are the one that are being used in industry and research sectors for varied applications, like growth rate determination, metabolic activity studies, protein purification etc.
Log phase cells have steady growth rate, hence it is easy to estimate the generation time (g) of the cells. Generation time is calculated as the time taken for the bacterial cells to double in their number. The log of cell number can be plotted against time (hours or days) to generate a predictable slope. The number of cells after certain period of time can be calculated from the formula:
N = Final cell concentration (or number)
N0 = Initial cell concentration
n = number of generations that happened over a period of time
When we apply log to the above formula,
Log N=Log No+n log102
n = 3.3(Log10 N-Log10 No)
The above formula can be used to calculate number of generations during the bacterial growth.
Generation time (g) can also be represented as t/n, where ‘t’ is the specific period of time in minutes, hours, days etc. With the amount of ‘t’ used during the growth starting from the initial cell concentration, ‘g’ can be calculated.
Stationary phase: When the nutrients get deprived and waste products/secondary metabolites start accumulating in the medium, bacterial cells are unable to further grow and enter into stationary phase. During this phase, number of cells being died will be more than the number of cells being produced. As a result, flattening of the growth curve happens at this phase. The new cells being produced also vary in its shape – from bacilli to more circular, size – large to small, cells- individual to aggregated formation etc because of the new starvation conditions that started prevailing in the growth medium.
All the physiological changes allow cells to survive the harsh environment rather than rapidly dividing. Cells also start producing secondary metabolites into the surrounding medium. Some bacteria like Bacillus form endospores and enter dormancy.
Decline/Death phase: Cells start dying off rapidly in this phase, resulting in the steepness in the growth curve. Most cells are irreversibly damaged that when transferred to fresh growth medium, very few cells may respond and show further growth. Some cells that are not able to revive their growth are called as viable but nonculturable (VBNC).
Bacterial growth kinetics
Optical density is a type of measure of Specific growth rate (µ). Spectrophotometer is used to measure optical density.
µ is calculated per hour as the generation time (T) at the time of lag phase as follows:
µ = (lnXn+1 – lnXn)/(tn+1 – tn)
T = 0.69/ µ
InX = natural log of OD
T = time of OD measurement
Bacterial growth is an autocatalytic reaction i.e., the growth rate is directly associated to cell concentration.
The equation of bacterial growth kinetics is represented as follows:
ΣS + X → ΣP + nX
Substrates + cells → extracellular products a+ more cells.
S = substrate concentration measured in gram per litre (g/L)
X = Cell mass concentration (g/L)
P = Product concentration (g/L)
n = increased number of cells or biomass
Net specific growth rate (1/time) is represented as
μnet = 1/X. dX/dt
t = time
Monads equation: Monads equation describes the dependence of the bacterial growth rate on the concentration of the substrate, as follows:
μ = μms / S +Ks
μ = specific growth rate of microorganism
μm = maximal specific growth rate
S = concentration of the substrate that is limiting source for growth
Ks = “half-velocity constant” (is the value of S when μ/μmax = 0.5)
Measurement of bacterial growth:
Following are the three major modes of measuring bacterial growth:
Microscopic count – counting the number of cells using a haemocytometer (originally used to count red and white blood cells). Haemocytometer is a special microscopic slide to count the cells in the gird present at the centre. However, it is not possible to distinguish live and dead cells unless cell permeable dyes like trypan blue is used.
Plate count method – A bacterial culture is serially diluted and spreaded onto the plate. After incubation for growth, colonies are counted which are direct representation of growth.
Turbidimetric method: Bacterial growth was measured based on the turbidity or cloudiness of the culture using a colorimeter. The number of cells grown is calculated by plotting the colorimetric values against standard graph generated using known cell numbers.
Apart from the above methods, bacterial growth can also be measured based on the dry weight and metabolic activity of the cells.
For microorganisms, soils are widespread and essential ecosystems that play crucial roles in providing plants with nutrients. If you dig a hole in the earth, you will find that the ground has a structure with various levels of evidence. These include the top organic horizon (O horizon), which includes freshly fallen litter on top by partially decomposed organic matter lower down, followed by horizon A, which comprises a range of minerals. The horizon B where humus, clays, and other materials transported live, and finally, the weathered parent material horizon C. With thin soils overlying calcareous regions of the world, such as the Yucatan, and dense soils occurring in some of the rich farmlands, such as those found in the Midwestern United States, the depth of these layers can differ drastically. Areas where heavy rainfall occurs, have nutrient-poor soils, such as the tropics, and over time the rains leach nutrients from the soils. The degree to which nutrients and microorganisms can travel can influence the permeability of the soil. Light does not penetrate beneath the first centimetre or two, eliminating phototrophy as a means of energy acquisition. The rhizosphere, the region around plant roots, is a habitat where abundant microbial populations occur.
Microbial Food Webs:
One of the greatest treasures of novel species is antibiotics and insights into how populations are organized in soil ecosystems on earth. On average, there are 109 bacterial cells in one gram of soil representing up to 5000 [or even 10,000] bacterial species by some estimates. The nature of soil food web and its inhabitants are more complicated than you might think. Plants play a significant role in physically structuring the under-ground ecosystem through their roots and the impact of their aboveground canopy, depending on the aridity of the aboveground environment. The larger organisms in the soil, such as earthworms, mites, springtails, nematodes, protists, and other invertebrates, were the subject of much of the soil biota research. These species help ‘engineer’ the soil conditions through their absorption and excretion of soil sections. On the scale of the microenvironment, bacteria and fungi even serve as engineers.
Soil microbial abundance varies with the microenvironment’s physical and chemical characteristics, including the content of moisture, organic matter abundance, and the size of soil aggregates. Although seasonal changes contribute to the dynamics of microbial soil populations, discussions of soil microorganisms typically refer to the topsoil sampled during the growing season. The most abundant microorganisms in the soil follow this sequence, as determined by plate count methods:
Microorganisms are most common on the surface and decrease as the depth increases in colony-forming units. In the variety of physiological forms of bacteria in various soil environments, soil types, including different organic content and related microbial processes, are seen. There is a substantial genetic diversity of bacteria in soil, and many of the physiological classes still have to be cultivated in the laboratory. Thus, molecular techniques produce more knowledge than conventional plating exercises for the study of the soil culture.
Soils comprise almost all the main microbial groups: bacteria, viruses, fungi, and archaea. Progress has been made in delineating which groups are most prevalent using culture-independent approaches in soil societies. Thirty-two different libraries of sequences from different soils were studied. Their results are impressive: 32 different phyla were present in the tests, but nine phyla were dominant: Proteobacteria, Acidobacteria, Actinobacteria, Verrucomicrobia, Bacteroidetes, Chloroflexi, Planctomycetes, Gemmatimonadetes, and Firmicutes. Proteobacteria account for the highest percentage of soils (39 percent on average). The majority of the sequences were new, and the results of this study vary significantly from the cultivation studies found in previous decades.
Symbiotic nitrogen fixers and mycorrhizae, which provide 5-20 percent of grassland and savannah nitrogen and 80 percent of nitrogen in temperate and boreal forests, are two main classes of bacteria in soil have been extensively studied. Nitrogen and phosphorus derived from symbiotic microorganisms are dependent on at least 20,000 plant species. Plants require nitrogen, and they are unable to fix atmospheric nitrogen in a beneficial form without their symbiotic partners. Examples of essential nitrogen-fixing bacteria are Frankia, an actinomycete that is important in forest growth, and Rhizobium, a key player in the health of crop legumes. These trees may grow in more marginal areas where nitrogen is restricted by interactions between plants, such as the alder tree (Alnus) and Frankia. More than 25 different genera of trees and shrubs have been recorded for cultivation in association with Frankia. Chemoheterotrophs, free-living in the soil or associated with a broad range of legumes, including alfalfa, clover, lupines, and soybeans, are rhizobia, including Azorhizobium, Bradyrhizobium, and Rhizobium. In order to infect them, plants release chemical compounds to attract soil rhizobia. On root hairs, most of the nodules form, but some form on stems. Nodules can use 7-12 percent of the plant’s photosynthetic production when active, but the expense is well worth the return in the form of fixed nitrogen available to the plant.
Despite the progress in understanding soil food webs, our understanding of soil food webs’ mechanisms is hindered by significant challenges. Different feeding classes are generally aggregated because, in their feeding patterns, most soil species are very “flexible,” muddling the distinctions between trophic stages. Relatively unknown are the diets of tiny species. New molecular techniques such as fluorescent in situ hybridization are exciting instruments that can help to expose the dynamic relationships in the soil population of who eats whom. Another instrument that is used to expose feeding relationships and energy sources is stable-isotope analysis.
For a human being who is 890,000 times larger than an E.coli cell, it is difficult to think of microbial environments on the order of micrometres to thousands of meters. Conditions like oxygen or pH will drastically change over this time. It creates microenterprises, and ecosystems are therefore more patchy than stable. Different abiotic factors influence and help establish these microenvironments in microbial communities in these habitats. Any disruption can lead over time to changes in microbial populations in habitats.
Table: Effects of abiotic factors:
Range of States
The ‘niche’ turning to the general ecological literature shows that there is what is known as the ‘fundamental niche,’ which reflects all environmental factors. In the ecosystems, the environment affects a species’ ability to survive and reproduce in the environment. The ‘realized niche’ is also the proper niche when biotic interactions (i.e., competition) restrict a species’ growth and reproduction. The definition of niche has been extended to microscopic organisms, enabling bacteria and archaea to exploit new niches not available to parentage by acquiring new genes through horizontal transmission. This niche definition for bacteria and archaea focuses more on the organism’s acquisition of new functional ability by transmitting horizontal genes, which implies a more complex character for niche boundaries. The survival and best reproduction niche of Ferroplasma are characterized by acidic, stable, rich in iron and heavy metals, and moderate temperatures. These conditions distinguish Ferroplasma’s niche space. Species also may change their climate so that other species have a more or less habitable environment.
The oceans and flowing water bodies, for example, rivers and streams, range in aquatic ecosystems. Water, more than 97% found in the world’s oceans, covers nearly 71 percent of the earth’s surface. In streams, rivers, and lakes, less than 1 percent of water is contained. Water is continuously renewed through the hydrological cycle in all those various marine ecosystems. The scale of aquatic environments and their diversity suggests the significance for microorganisms of aquatic habitats. Key microbial players in aquatic environments include primary production phototrophs and the heterotrophs involved in carbon cycling in aquatic habitats.
In these marine settings, the environmental and physicochemical conditions vary greatly. Water movement is one of the apparent factors; streams and rivers will flow quickly, with lakes moving less. Winds produce surface water movement in the seas, create ocean waves, and create upwelling areas. These winds transfer nutrients, organisms, oxygen, and heat worldwide, in addition to deep-water currents. As in the seas, water circulation in all marine environments determines different properties of water. Physicochemical factors, such as the pH, the abundance and availability of macro-and micronutrients, salinity, phosphorus, nitrogen, sulphur, and carbon, can vary significantly within the various ecosystems.
Table: Characteristics of different aquatic habitats:
-1.5 to 27oC at surface
Lakes a. freshwater b. great salt lake
0.01 avg. 12%
Storms are very fluid, have significant variations in physical and chemical environments, are greatly affected by their drainage range, and have a single water flow. In contrast, the lakes, particularly the stream’s headwaters, have more stable conditions and primary productivity. Lakes can be acidic or alkaline (e.g., Mono Lake, California), but often they can be saltier than freshwater, like the Great Salt Lake, Utah.
Aquatic microbial ecology has been advanced from descriptive research on who’s home” to hypothesis-driven studies of interactions and environmental and biological controls on the diversity and population distributions. A broad range of anti-predating mechanisms, including the secretion of exopolymer substances and capsules made up of polysaccharides and morphologic adaptations, are two of the exciting features and the subject of several studies. They are gram-positive. Studies have focused on how predators evade microbial communities, like predation, particularly by protists, and virus lysis is a significant mortality factor. Viruses in various aquatic environments are standard, with a difference of between one or two orders in size in these different habitats, while in freshwater, the abundance of the virus is more seasonal. In aquatic settings, what governs viral abundance is still under review. In aquatic environments, viruses have a vital role in recovering organic matter dissolved by lysing their presence into their bodies, converting the carbon and other nutrients.
a. Fresh Water
The word wetlands for freshwater generally applies to rivers, streams, reservoirs, lakes, and groundwater. Freshwater that contains less than 1.000 mg/ l dissolved solids is classified under the United States Geological Survey (USGS). As noted above, freshwater microorganisms vary greatly from marine environments in their phylogenetic diversity. Typical freshwater bacterial classes include beta-proteobacteria (e.g., the relative of Rhodoferax and Polynucleobacter necessarius), Actinobacteria, Cytophaga/Flexibacter/Flavobacterium hydrolysis relatives.
Lakes are aquatic lakes, initially formed by glaciation, volcanism, or tectonics. The Great Lakes in North America, and Lake Baikal, Siberia, comprise approximately 40% of the world’s freshwater at a few vast lakes.
There are many gradients within water bodies that affect microbial distribution populations. The oxygen gradient is one of the most critical. In lakes where upper waters can be oxic and warmer, the lower gradient is colder and often anoxic. The thermocline is separated by these two layers, which is a transition region between the two layers. Seasonal changes in atmospheric temperature and water temperature can result in changes in density that turn the water over and allow oxygenated water to enter the lake’s lower reaches. It influences the microbial communities of the lake.
The vegetation around lakes supplies some nutrients that have been found in lakes. Low nutrient quantity lakes are oligotrophic, while high nutrient quantities, productivity, and oxygen depletion can affect species that can survive under such conditions. Lakes are eutrophic. The transformation of contaminants such as sulphur dioxide and nitrous oxide (NO3) into acid rain causes some lakes to be naturally acidic while other acidic. Lakes in North-East America recover from acid rain impacts. The pH of the water in lake also influences the population of microbes.
c. Rivers and Streams:
During and after a rainstorm, the water will change and get a water movement force in rivières. Water is moving in streams and rivers through vast material, soil, trees, rocks, and other substances. It ensures a steady supply of nutrients to biotic communities and a great deal of trouble during floods. Many rivers cross cities and thus are exposed to human wastewater and other contaminants that can directly affect the river’s population. As the metabolic diversity of microorganisms is such that specific contaminants are potential energy sources in microorganisms. Since high organic loading can result in high productivity that diminishes oxygen levels, areas of urban rivers can be anoxic, limiting microorganisms in such regions.
The ecosystem of the river consists of many components like horizontal
(1) the active channel that can go dry part of the year in some rivers and streams and
(2) the transitional zone between the marine and the terrestrial habitats, in the riparian zone.
Vertically, streams and rivers are marked by
(1) Waters of the surface;
(2) the sub-surface water region of the hyporheic zone;
(3) the phreatic groundwater field.
The physicochemical properties of these ecosystems differ. Rivers and streams have many suspended organic and inorganic particles, restricting how much light penetrates the water column. At least partly shaded by trees that hang over the streams, the parts of the reaches have extensive vegetation. The extent of photosynthesis in the streams is restricted by both turbidity and shading. Desert streams are much higher than in tropical and temperate regions and have no shading of microbial photosynthesis. Rivers and rivers differ in their salinity by order of magnitude; desert rivers have the highest amounts.
d. Hot Springs:
Springs are springs of geothermal water, groundwater which comes into contact with hot rocks or magma from the world’s earth’s crust in volcanically active regions. Some impressive examples are found in Yellowstone’s national park in Wyoming, Iceland, Japan, and New Zealand. Hot springs reflect extreme temperature conditions and, in some cases, pH. There is a high concentration of anaerobic or microaerophilic hot spring that suggests low oxygen concentrations. In hot springs where temperature limits are photosynthesis, they were suggested to be primordial producers. Hyperthermophiles, who use carbon dioxide as their carbon source, are also chemoautotrophs and serve as primary producers within hot spring ecosystems. Hot springs contain various gases, including molecular hydrogen and a reduced number of iron and sulphur compounds, dissolved and provide electron donors. It implies that Yellowstone’s primary productivity results from molecular hydrogen oxidation, which can happen to levels above 300 nM. in hot springs. Hot springs are the prime habitat for archaeological animals.
In addition to the fact that it is a saline ocean rather than a terrestrial habitat. It is one of several environmental parameters that influence the existence of marine habitat microorganisms. Furthermore, temperature, light, food supply, and pressure vary from the surface to the ocean’s depths. The ecosystems of the ocean shift from shore to vertical depth. Traveling further into the water, from the surface or epipelagic region to the mesopelagic zone (200–1000m), you move into the bathypelagic zone (1000–4000m), the abyss area (4000–6000m), and eventually the Hadean area (<6000m).
b. Food and Microbial Aquatic Habitats:
The marine food web is typically characterized by the low nutrient abundance and patchy nature of the gradients, as mentioned above, and high salinity.
Although the food web has been studied on the ocean for more than a hundred years, several recent findings have led us to believe that the classical description of a chain from diatoms through copepods and to fish and whales can only be a small part of the energy flow. Recent studies of microorganisms, organic dissolved matter, and organic particles in the sea have shown other mechanisms by which a significant share of the energy available will flow. For decades, marine scientists have been cautiously approaching this food web view, and care should be taken when a paradigm is challenged.