Methyl Red – Voges Proskauer (MRVP) tests

 

The methyl red (MR) and the Voges-Proskauer (VP) tests are used to differentiate two major types of facultative anaerobic enteric bacteria that produce large amounts of acid and those that produce the neutral product acetoin as end product.  MRVP tests are of value in the separation of Escherichia coli and Enterobacter aerogenes, both coliform bacteria, which appear virtually identical except for certain physiological differences that are used as indicators of the sanitary quality of water, foods, food-production and eating establishments.  Both these tests are performed simultaneously because they are physiologically related and are performed on the same medium MRVP broth. Opposite results are usually obtained for the MR and VP tests, ie., MR+, VP^_    or MR^_, VP+.

Aim

To find out whether an organism is MR-VP positive or negative.

Principle

The hexose monosaccharide glucose is the major substrate oxidized by all enteric organisms for energy production.  The end products of this process will vary depending on the specific enzymatic pathways present in the bacteria. In MR test the pH indicator methyl red detects the presence of large concentrations of acid end products.  Although all enteric microorganisms ferment glucose with the production of organic acids, this test is of value in the separation of    E. coli and E.  aerogenes.

Both of these organisms initially produce organic acid end products during the early incubation period.  The low acidic pH (4) is stabilized and maintained by E. coli at the end of incubation.  During the later incubation period, E.  aerogenes enzymatically converts these acids to nonacidic end products such as 2,3-butanediol and acetoin (acetyl methyl carbinol), resulting in the elevated pH of approximately 6.  The methyl red indicator in the pH range of 4 will remain red, which is indicative of a positive test.  At a pH of 6, still indicating the presence of acid but with a lower hydrogen ion concentration, the indicator turns yellow and is a negative test (Figure 1).

The VP test determines the capability of some organism to produce non acidic or neutral end products, such as acetyl methyl carbinol, from organic acids that result from glucose metabolism, which is characteristic of E.  aerogenes (Figure 2).  The reagent used in this test, Barritt’s reagent, consists of a mixture of alcoholic α-naphthol and 40% potassium hydroxide solution.  Detection of acetyl methyl carbinol requires this end product to be oxidized to a diacetyl compound.  This reaction will occur in the presence of the α-naphthol catalyst and a guanidine that is present in the peptone of the MRVP medium.  As a result, a pink complex is formed, imparting a rose color to the medium.  Development of a deep rose color in culture 15 minutes following the addition of  Barritt’s reagent is indicative of the presence of  acetyl methyl carbinol and represents a positive result.  The absence of rose coloration is a negative result (Figure 3).

Requirements

24 hr nutrient broth cultures of species Bacillus, Streptococcus, Staphylococcus, Pseudomonas and E.coli.

MR-VP broth (5ml / tube), Methyl red indicator, Barritt’s reagent A and B, Bunsen burner, inoculating loop, glass marking pencil, dropper etc.

Procedure

1.  Divide the total MR-VP tubes into two sets.  Using sterile techniques, inoculate each set of   MR-VP broth with given cultures of organisms.  Uninoculated tubes serve as control.

2.  Incubate the tubes at 37^o C for 24-48 hrs.

3.  Add 5 drops of methyl red indicator to the first set of organisms and observe the color change for MR test.

4.  To the other set of tubes, add 12 drops of Barritt’s reagent A and 2-3 drops of Barritt’s reagent B.  Shake the tubes gently for 30 seconds with the caps off to expose the media to oxygen.

5.  Allow the reaction to complete for 15-30 min and observe the color change for VP test.

Observations

In the MR test, the MR-VP broth remained red color in the tubes inoculated with Bacillus, Streptococcus, Staphylococcus and E.coli. No change observed in tubes inoculated with  Pseudomonas sp.

In the VP test, no development of a crimson to ruby red color formed in any of the tubes inoculated.

 

Result

Among the given culture of organisms Bacillus, Streptococcus, Staphylococcus and E.coli are MP positive and VP negative whereas  Pseudomonas sp. is MR and VP negative.  

       Barritt’s reagent A and B

 Solution A

Alpha-naphthol                                   5.0 g

Absolute ethanol                                 95.0 ml

(Note: Dissolve the alpha- naphthol in the ethanol with constant stirring).

Solution B

Potassium hydroxide                          40.0 g

Creatine                                               0.3 g

Distilled water                                    100 ml

(Note: Dissolve the potassium hydroxide in 75 ml distilled water. The solution will become warm. Allow to cool to room temperature. Add the creatine and stir to dissolve. Add the remaining water. Store in a refrigerator). 

MR-VP broth                                            

Peptone                                               7.0 g

Dextrose                                              5.0 g

Potassium phosphate                           5.0 g

Distilled water                                    1 litre

pH                                                       6.9

International Standard Serial Number (ISSN)

 

ISSN is an 8-digit identifier code assigned for regular publications like newspapers, journals, magazines and periodicals of all kinds including electronic publications.

1.    What is ISSN?

Authors always prefer to publish their articles in periodicals with ISSN. ISSN means International Standard Serial Number which is a unique international identifier for serial publications. It is universally accepted as a means to identify serial publications and are assigned to journals since 1970s.  From the end of  1990s, ISSN is also assigned to electronic serials (online, CD-ROM, DVD etc.) and to electronic “ongoing integrating resources” like websites and databases. Thus ISSN formed an eight-digit number which is an identification for periodical publications, including electronic serials. The ISSN is a numeric code which is used as an identifier. The ISSN number is published as -the acronym ISSN followed by two groups of four digits separated by a hyphen. The eighth character is a control digit calculated according to a modulo eleven algorithm on the basis of this seven preceding digits. This eighth control character may be an “X” if the result of the computing is equal to “10”, in order to avoid any ambiguity (figure 1).

 

Fig 1- ISSN Number

2.    Importance of ISSN

ISSN number and copyright of the journal are not at all related and the procedure to obtain both are entirely different. ISSN is generally printed on a prominent place, such as at the top right hand corner of the front cover, near the title on every issue. The number of copies produced is not related to the ISSN assignment. ISSN is assigned to publications that are intended to be published continuously on a regular or irregular basis. Eg: Journals, Proceedings, Annual reports, Quarterly reports, Biannual reports, Bulletins, Newsletters, proceedings etc. The ISSN identifies the title of the serial publications and the same ISSN can be used as long as the title remains the same. If there is any change in the title, it is mandatory to inform the ISSN office. Title change would also cause change in ISSN. ISSN is also assigned to good quality ‘Online publications”. Also, other types of regular electronic publication formats, such as DVD’s and CD-ROM’s are assigned with ISSN. If same periodical is published in different languages, then separate ISSN is required for each language. There is no expiry date for ISSN. If the publisher is unable to publish the forthcoming 2 issues, then also it should be intimated to the ISSN office.

3.    How to get ISSN?

Each country is having a respective national authority for ISSN. Publishers can apply for ISSN in advance of publication. Application can be completed and submitted online or can be sent in printed format through post or fax to the ISSN international authority for international publication and ISSN national authority. All details can be had from the ISSN website http://www.issn.org. Basic informations required to apply for ISSN includes the proposed title of publication, frequency of publication, proposed start date (month / year) and publisher name and address. However, first issue of publication has to be sent to the centre in order to verify and validate the details. This copy will be passed through to the Legal Deposit Office and subsequent issues can be sent directly to them.

ISSN can be also obtained for existing serial publications. Application for that should include all above mentioned details and some additional informations and documents as mentioned below.

·         For printed publications –A copy of  recent issue

·         For on-line journals- Either the URL or any passwords required (as requested on the form); or a printout of the title screen, and any screens giving publisher information (company name and the place of publication).

·         For CD-ROM or diskette journals- a sample copy or a printout of the title screen together with copies of the labelling, documentation and packaging.

Sample ISSN numbers of the publications

ISSN 0027-9633 (online)

ISSN 0027-9634 (print)

Reference

Achalare, R. A., Patil, S. V., Patil, S. S. 2014. Significance of ISSN and ISBN in Publications, Journal of Current Pharma Research, 5(1), 1378-1381.

 

 

 

 

Air borne bacterial diseases

A number of diseases are transmitted through air. Such diseases are called airborne diseases. Airborne diseases are divided into bacterial, viral and fungal diseases. Microorganisms released as droplets, Droplet nuclei,Infectious dust and spores are easily disseminated through air. Airborne bacterial diseases are mainly respiratory tract infections transmitted by droplet inhalation or by contact with contaminated inanimate objects. Upper respiratory tract consists of nostrils, pharynx and tonsils. Lower respiratory tract consists of larynx, trachea, bronchi, brachiolus, alveoli and lungs. Transmission of fungal diseases is more through air. But the airborne bacterial diseases are more in number. Common airborne bacterial diseases are as follows.

1.        Brucellosis

2.        Meningococcal meningitis

3.        Meningitis by Haemophilus influenzae

4.        Pulmonary Anthrax 

5.        Diseases by Streptococcus

6.        Atypical pneumonia

7.        Psittacosis

8.        Diphtheria

9.        Pertussis (Whooping cough)

10.    Tuberculosis

11.    Legionellosis

12.    Tularemia

 1. Brucellosis

Brucellosis is also known as “undulant fever” because of its characteristic raising and falling nature of temperature.Brucellosis is a zoonotic disease caused by Brucella species. Brucella species includes B. abortus, B. melitensis,B. suis and B. canis. They are tiny, faintly staining Gram Negative coccobacilli. Brucellosis is a disease of humans, animals and birds, mainly caused by  B. suis, commonly occurring in infected swine. Inhalation of aerosol during processing of infected swine causes infection in humans. Infection is commonly found in veterinarians, butchers and slaughterhouse workers. Hence it is considered as an occupational disease. In humans the disease is characterized by prolonged and undulating fever, headache, chill, generalised aches and pains of muscles and joints. Mortality rate is low, less than 2%.

 2. Meningococcal meningitis

Meningitis is the inflammation of meninges. Meninges is the three layered membrane covering the spinal cord and brain. The term meningitis is derived from a Greek word. “Meninx” means  Brain and “itis” means inflammation. This is  considered as a potentially serious disease as the microbial invasion is to the nervous system. Meningitis is divided into bacterial or septic meningitis and aseptic meningitis. Meningococcal meningitisIs a fatal septic meningitis caused by Neisseria meningitidis. Neisseria meningitidis Is a gram-negative diplococci.

About 10% of population are healthy carriers and harbour this bacteria in the back of the nose and throat. Healthy carriers, Convalescent carriers and infected persons release this cocci as respiratory droplets by activities like coughing, sneezing, kissing, talking etc. Disease is also transmitted through sharing of utensils. No animal reservoir is found for this disease. Inhalation of these bioaerosols will lead to the infection

Organism first colonize in the nasopharynx and penetrate the mucosal barrier to reach the blood stream. They cross the blood-brain barrier and enter cerebrospinal fluid (CSF) and cause meningitis.

 3.  Meningitis by H. influenzae

Mode of infection and disease are the same as above. But the infection is commonly found in children between 6 weeks and 2 years of age. The causative agent is Haemophilus influenzae which is a Gram Negative coccobacilli. Infection can also cause secondary pneumonia in patients.

 4. Pulmonary Anthrax

This is also called ‘Wool sorters disease”. Pulmonary anthrax is mainly a disease of herbivorous animals. It is a highly infectious animal disease that can be transmitted to humans by direct contact with infected animals like a cattle,goat, sheep etc or through their products.

Anthrax is of three types. Human infections through cut or abrasion of skin lead to cutaneous anthrax.   If spores reach gastrointestinal tract, it cause gastrointestinal anthrax. Inhalation of the spores leads to pulmonary anthrax or wool sorters disease. Pulmonary anthrax is seen only in human beings. Causative agent is Bacillus anthracis which is large, Gram Positive, aerobic, endospore forming bacteria. It is world wide in its distribution. Disease transmission is by inhalation of the dust contaminated by animal products. Usually through the sorting or combing of raw wool contaminated with spores. Spores are usually 1 to 2 to µm in diameter and  can enter the lower respiratory tract easily and lodge in the alveolar spaces. There they are engulfed by alveolar macrophages. But can resist and survive the phagocytosis, germinate within the endosome. Bacteria then spread to regional lymph nodes and eventually reach the bloodstream. This will result in pulmonary anthrax which is characterized by massive pulmonary edema, hemorrhage and respiratory arrest.

 5.  Diseases by Streptococcus

A number of airborne bacterial diseases are caused by Streptococcus species. Streptococci  are large group of Gram Positive cocci and most significant one causing air borne infections is S.pyogenes. Infection is either due to the multiplication of pathogen or due to the toxin produced by the organism. Infection may range from mild diseases like most  common “Strep throat” to life threatening conditions. Organisms reside in the nose and throat and are transmitted through respiratory droplets.

a) Streptococcal pharyngitis

This disease is commonly called Strep throat or tonsillitis or sore throat. It is a common mild infection predominantly found in children of 5 to 15 years of age. Symptoms include fever, headache, red and sore throat, enlargement of lymph nodes of neck. Often results in bleeding throat.

b) Scarlet fever

Sometimes Streptococcal pharyngitis proceeds to Scarlet fever. This condition is due to the production of “erythrogenic toxin”  by Streptococci. The disease characterized by strawberry tongue and red skin rashes.

c) Rheumatic fever

3% of untreated upper respiratory tract infections by Streptococcus pyogenes and repeated early childhood infections may lead to a condition known as Rheumatic fever. This disease is characterized by inflammation and degeneration of heart valves and joints.

d) Streptococcal pneumonia 

About 60-80 % of all respiratory diseases known as pneumonia are caused by Streptococcus pneumoniae. Infection occurs in those individuals with predisposing factors such as viral infections of respiratory tract like common cold, or physical injury to the respiratory tract, alcoholism, or diabetics. Causative agent Streptococcus pneumoniae is a gram-positive organism, normally found in the upper respiratory tract. Virulence factor of this organism is the capsular polysaccharide composed of hyaluronic acid. Capsular polysaccharide protects the organism from ingestion and killing by phagocytes. Thus bacteria will multiply in alveolar spaces and also produce toxins called “pneumolysin” which may destroy the host cells. Alveoli get filled with blood cells, fluids and become inflamed. Lung inflammation is a characteristic feature of this disease. Sputum is often rust-colored due to the blood coughed up from lungs. Onset of the disease is a abrupt with chills, hard laboured breathing and chest pain. Inhalation of aerosols liberated from the infected person leads to infection.

 6. Atypical pneumonia

 Is an infection of lower respiratory tract  and also called “walking pneumonia”. Causative agent is Mycoplasma pneumoniae. Symptoms are gradual in onset and are milder than typical Pneumonia. Symptoms include  persistent cough, headache, fever, sore throat, chest pain while deep breathing etc. Cough, sneeze etc will liberate bioaerosols from the patient and inhalation of these will lead to infection.

 7. Psittacosis

This infection is mainly acquired from birds. Infection may range between mild respiratory diseases to severe pneumonia. Infection may sometimes lead to encephalitis, coma and convulsions. Sometimes fatal infections may occur. Causative agent is Chlamydia psittaci. Pathogen is liberated as infectious dust from the infected avian faeces. Inhalation of the infectious dust will cause disease in humans. Hence this is considered as an occupational disease of poultry workers and handlers.

 8. Diphtheria

Causative agent is Corynebacterium diphtheriae which is a gram positive bacilli. Is mainly found in children. Toxins produced by these microbes cause generalized toxemia in patients. Infection occurs in the upper part of the respiratory tract and nose, throat and tonsils get inflamed.  Also lymph glands in the neck region swell and result in “bull neck appearance”. Powerful exotoxin produced by the pathogen destroys the cells cof the epithelial lining. Thus  the dead cells, mucus and scavenger cells pile up and form a pseudo membrane. This membrane is leathery in consistency and cause respiratory blockage and death by suffocation, particularly in young children. Tracheotomy (Cutting hole in throat) are performed to save children. Sometimes, toxin diffuses to blood stream, causing widespread damage particularly to the heart.

 9. Pertussis (Whooping cough)

Is highly infectious and potentially lethal disease caused by Bordetella pertussis which is a Gram negative coccobacilli. This disease mainly affects infants and young children below 4 years of age. No non human reservoir for this disease and infected children act as source of infection. From infected person the pathogen is liberated as aerosols through activities like talking, coughing, sneezing, laughing etc. Bacilli binds to ciliated epithelium of upper respiratory tract and secrete toxins. The function of toxin is to clear mucus from air passages and hence they damage cells. Net effect is the build up of stick glue like mucus in airways. This will result in common cold like symptoms followed by spasms of violent, hacking, persistent, recurrent cough with at least 15 to 20 coughs at a time. This is actually an attempt to remove the accumulated mucus from the respiratory tract. But these episodes result in oxygen deficiency, triggering deep and rapid inspirations through the partially obstructed air passages, resulting in characteristic ‘whoop’. Coughing may be so violent that it may cause vomiting,  hemorrhage and even brain damage. It may last for several weeks or months.

 10.  Tuberculosis

Tuberculosis or TB is a severe lower respiratory tract disease. Causative agent is Mycobacterium tuberculosis which is an acid fast bacteria. These bacteria have high mycolic acid content in the cell wall which makes a waxy coating on its surface and organisms are seen as clumps. These are highly virulent bacteria and even a single organism is  sufficient to initiate an infection. In a susceptible host, the inhalation of a single particle carrying a viable tubercle bacillus and  small enough to reach the lungs is capable of producing infection. Disease is characterized by loss of appetite, fatigue, weight loss, night sweats and persistent cough. Inhalation of aerosols liberated from the patient is a mode of infection.

 11. Legionellosis

This is also a lower respiratory tract infection. This disease is considered as a type of bronchopneumonia or atypical pneumonia. Causative agents are coming under Legionella species.  Legionella species Are Gram Negative bacteria found naturally in fresh water bodies. They may contaminate Air conditioners, cooling towers or showers and cause infection. No person to person transmission of disease occurs.  Legionellosis is of two types. Pontiac fever and Legionnaires disease. 

Pontiac fever is a self limiting, non-fatal disease. It is caused mainly by Legionella  pneumophila. Symptoms include fever, chills, dry cough and headache  with mild upper respiratory tract infections resembling acute influenza. This resolves spontaneously and often goes undiagnosed. Inhalation of mist from water sources like air conditioners, cooling towers, showers etc., contaminated with the pathogen is mode of infection.

Legionnaires disease, unlike Pontiac fever, is severe bronchopneumonia and can be fatal. Here also the causative agent is Legionella  pneumophila. Symptoms include fever, chills, dry cough, vomiting and diarrhoea.  Pathogen occurs in natural air and in freshwater. At times they enter and proliferate in cooling towers, air coolers and showers. Praying and splashing of water containing pathogens may produce aerosols which are disseminated in air and inhalation causes infection. 

 12.  Tularemia

Causative agent of the disease is  Francisella tularensis, which is a Gram negative bacilli. This is one of the most virulent bacteria, inhalation of as few as 10 organisms is sufficient to establish the disease. It is also known as rabbit fever as it is a zoonotic disease transmitted from natural reservoirs of small mammals  like rabbits, mice and squirrels. Here also, there is no transmission of disease from person to person. Inhalation of dust particles liberated from the infected animals result in severe respiratory illness leading to pneumonia. 

Selection of Journals for Publication

 

Today, around 30,000  peer reviewed journals are there, publishing approximately 2 million articles every year. It is very important to find out the apt journal for your scientific work. Selection of journal for publication of our research work is a difficult task. An author should look into a number of factors before selecting a journal for publication. Relevance, acceptance rate, circulation, prestige, publication time etc are certain primary things to consider.  Selection of journals are also dependent on a number of other factors as mentioned below.

1.        Specialization spectrum & Target group of the Journal

Before preparing an article, authors should recognise the main theme and predominant message there are going to convey through the publication and can select a journal that is publishing articles in that specialised spectrum. Authors should also think about the target group to whom this message should reach. Depending on the target group, authors could make a list of journals that are primarily read by the target group. It would be also useful if the authors can find whether their message is meant for international, national or regional readers and thus can accordingly select the journals. Sending papers without doing such things may lead to the rejection of article.

On receiving an article, Journal Editor first checks whether the article is of interest to their target group or readers. So, after selecting a journal for publication, the author should first check the types of paper published in that particular journal. Author can also see the last few issues of the journal if available and could find out whether the journal has published papers similar to those which we intend to write.

2.        Peer Reviewed Journal

Journals which publish articles only after being evaluated by other experts in the field are called Peer Reviewed Journals.   Peer review is a multi-layered review process in which the article is evaluated by Editors, reviewers and statiticians. Editor would check whether the article meets the focus of the journal and is suitable.  Then the article undergoes review by Technical Editor who verifies whether the format of article adhere to the prescribed journal style. Then, Section Editors verify the contents. Statisticians would verify appropriateness of statistical methods used in the analysis of research data. Then the article moves to the Subject Experts or Reviewers in the particular field. Thus, the peer review is a time consuming process, however to get published in good quality journals, articles have to undergo this peer review process. Some journals easily publish papers without much review. Such publication are not valued highly by the scientific community.

3.        Acceptance Rate  

All authors wish to avoid rejections. So before submitting articles it would be better to check the acceptance rate of journal from its website. Journals that are not very prestigious have better acceptance rate. Acceptance rate is less for good journals. Usually all journals are interested in quality publications. Hence well written articles with novel idea, strong evidence, solid and meaningful conclusions would be considered for publication.

4.        Indexed Journal   

Multiple databases index journals on the basis of various criteria like quality, regularity of publication, professional standing, review process etc. Hence journals included in such databases are called “indexed journals” and are considered as prestigious. In many professions, promotion to higher posts makes it compulsory to have publications in indexed journals. PubMed is the largest database for biomedical journals all around the world. It is freely available on internet and most popular among scientists, researchers and academicians.  

5.        Impact factor

Impact Factor (IF) is one of the means to assess the value and importance of a journal. It is considered as the most popular assessment means for journal quality among authors, institutions and employers. It is calculated on the basis of number of citations received by journal articles in proportion to the number of citable article published over a period of time. IF in one year is calculated by dividing the number of citations to articles in the previous two years in that particular journal by number of articles published by that journal in the 2 preceding years.

For Eg: Suppose , you want to calculate the IF of Americal Journal of Pathology for the year 2020.

If,             Journal citations in 2019 =   130

Journal citations in 2018 =   170

and

Number of articles published in 2019 = 30

Number of articles published in 2020 = 25

Then, IF = 130 +170/ 30+ 25 = 300/55 = 5.45

IF of Journal in 2020 = 5.45

Usually IF cover last two years. Hence IF change from year to year. In resume, along with publications, researchers also give their impact factor. High impact factor journal publications are highly valued but rejection rate is more.

6.             Open Access Journals   

The journals which provide unfettered access to the entire research content they publish are called Open Access Journals. Articles published in such journals receive wide audience. But some such journals collect publication charge from authors.

7.    Journals Published by Professional Societies

Many Professional bodies and societies publish journals of their respective field and free copies are given to the members. This ensures that the journal is distributed widely among people in that particular field. Authors could publish in such journals if the authors wish to communicate their message to members within a particular specialty.

8.                  Publication in Print or Online version

Most of the journals are published as printed version and also have online version. Publishing in journals having both these versions is highly recommended.  Printed articles have high prestige whereas online journals increase the article access and author visibility.  Printed articles have been a great pride and pleasure for authors. But experts assume that the cost of printing and publishing can be reduced by online journals. Also it adds universal availability through inexpensive internet access and articles could be published rapidly. Hence, printed journals might become a rarity in future.  

Online journals publish articles in the web for everyone to read, so reach even those who have not subscribed the journal. Papers in Online journals have additional features that benefit readers. For example, sometimes links to other related papers are cited in reference.

9.             Cost of Publication

Some journals charge fee for publication. Fee could be collected at any stage before publication. It could be at the time of submission, review or once the article is accepted for publication. Some such journals provide fee concession for publication if study was done in resource poor settings or as part of unfunded projects, or if primary author is a student.

Some unpaid journals collect fee if the word count or the number of pages, figures etc exceed the prescribed limit. Some such journals charge for the coloured figures. Publishing in paid journals increases the cost of publication. Hence authors should first check fee details before selecting the journal for publication.

10.         Efficient Journal Administration

 Before selecting the journal author should see whether the journal is well managed and being published regularly. The journal should be author friendly and also should provide fair and time bound review process. Desirable attributes of author friendly journals are as follows:

·         Electronic submission facility

·         Prompt acknowledgement of receipt of  manuscript

·         Fair peer review process

·         Facility of tracking the article

·         Time bound review process & publication

· Responsive editors & transparent decision making process (Editor should give prompt answers to the queries and concerns of the author

·          Availability of additional services for manuscript. (Some journals give services of qualified statisticians and also service to people to improve language, whose native language is not English). 

·         Regularity of publication

References

1.      Bavdekar, Sandeep. (2015). Choosing the Right Journal for a Scientific Paper. The Journal of the Association of Physicians of India. 63. 56-59.

2.      James Hartely, 2008. Academic writing and publishing-A practical handbook (ISBN 0-203-92798-2), Routledge Taylor & Francis Group, New York