Modulation Of Antibiotics Resistance Pattern In Escherichia Coli By Different Plant
Material type: Book ; Literary form:
Publisher: 2015 Dissertation note: Escherichia coli (E. coli) is Gram negative microorganism belonging to family
Enterobacteriaceae. It is part of normal micro flora of gastrointestinal tract of human and all
warm blooded animals (Kaper et al. 2004). Escherichia coli is source of many infectious diseases
in human as well as in animals. Common E. coli infections are enteritis, urinary tract infection,
septicemia and neonatal meningitis. In pets and farm animals, E. coli is associated with diarrhea
(Allocati et al. 2013). Poultry industry is facing huge annual losses due to infection of avian
Pathogenic E. coli (APEC) in broilers (Oosterik et al. 2014). E. coli causes a variety of
syndromes in poultry including yolk sac infection, respiratory tract infection, swollen head
syndrome, septicemia and cellulitis (Buys et al. 1989)
Antibiotics are chemical agents which inhibit the microbial growth and used to eradicate
infections. Mechanisms of action of antibiotics provide a base to categorize antimicrobial agents.
Most important classes of antibiotics act as inhibitors of cell wall synthesis, protein synthesis
(tetracyclines and macrolides), nucleic acid synthesis (fluoroquinolones), metabolic pathway
(trimethoprim-sulfamethoxazole) and cell membrane (polymyxins). Bacteria may have intrinsic
or acquired resistance to antimicrobials (Tenover 2006).
Urinary tract infections are mostly caused by E.coli. Antibiotics generally used for the treatment
of E. coli infections include ampicillin, nitrofurntion, cephalosporin, sulphonamides
(trimethoprim-sulfamethoxazole) and quonolones (neladixic acid, ofloxacine, ciprofloxacin and
levofloxacine) (Lin and Lin 2010).
Extended use and misuse of antibiotics lead to the development of resistant bacteria. Resistant
E. coli strains are common source of hospital born and community acquired infections. Ease of
international travelling is one of the major spreading factor for antibiotic resistance. Resistant
bacteria got opportunity to move from one geographical area to another (van der Bij and Pitout
2012). New strains of E. coli resistant to carbapenems (New Delhi metallo-β-lactamase 1 (NDM-
1) are major global health issue (Kumarasamy et al. 2010). Antibiotic resistance has become a
serious public health problem. Currently, world is facing great difficulty in treatment of many
infectious disease of human and animals. One of the reasons of treatment failure is emergence of
resistant bacteria (Levy 2002).
To develop new strategies for treatment of infectious diseases, it is necessary to understand the
mechanisms of resistance. Efflux pump inhibitors, enzymatic degradations and alteration of
target sites are major strategies by which bacteria acquire or develop resistance to antibiotics
(Sibanda and Okoh 2007).
Scientists are looking for alternatives of antibiotics such as bacteriopheges, naturally
antimicrobial compounds and some non antimicrobial agents (Worthington and Melander 2013).
Probiotics (Lactobacillus and bifidobacterium) can be a prophylactic measures against E. coli
and may be used to treat intestinal tract infections of E. coli and other bacteria (de Vrese and
Phytochemicals, secondary metabolites of plants, have antibacterial activity against many
pathogenic organisms. These phytochemicals in combination with antibiotics may show
synergistic effect. Phytochemicals and plant extracts can be a source of antibiotic resistancemodifying
agents (RMAs) (Abreu et al. 2012). Plant extracts shown antibacterial activity
because of phytochemicals like alkaloids, tannins, flavonoids, phenolic compounds and steroids
(Gobalakrishnan et al. 2013). Plant extracts are used as traditional medicine for the treatment of
many diseases. Plant extracts like Zingiber officinalis (Ginger) Gymnema sylvestre (Gurmar buti), Astragalus (goat’s thorn), Calotropis procera (apple of Sodom) and oputia dillenii (cactus)
have antimicrobial activity (indu et al. 2006 and Kumaar et al. 2013). Plant extracts also have
antibiotic resistance modulation potential (Mako et al. 2012).
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Antimicrobial Potential Of Bovine Lactoferrin Against Foodborne Pathogens
Material type: Book ; Literary form:
Publisher: 2014 Dissertation note: Health is recourse of everyday life, but not the object to live. It is positive to give special importance to personal and social resources. However, in Pakistan and other developing countries conditions are different, most people have low income and they live in un-sanitized environment. They eat un-hygienic food and also lack safe drinking water. People do not adopt any preventive measure to minimize the risk of contamination. Food storage is also un-hygienic. These conditions lead towards contamination and result in foodborne infections and gastro-enteritis. Foodborne illnesses are always a serious health issue in the Pakistan and throughout the world. Individual’s record for foodborne illnesses is impossible but it is reported that 7 out of 10 people suffer from foodborne illness caused by different microbes each year worldwide (WHO survey 2012).
Foodborne illness is caused by eating contaminated food with pathogenic bacteria. Some common pathogens are Escherichia coli, Campylobacter jejuni, Clostridium botulinum, Clostridium perfringens, Listeria monocytogens, Bacillus cereus, Staphylococcus aureus and Salmonellaspecies. Incubation period for onset of symptoms of food poisoning ranges from hours to days. Nausea, vomiting, abdominal cramps, fever and diarrhea are symptoms which appear commonly in most of food poisoning. However, foodborne illnesses if left untreated can lead severe dehydration, imbalance of intestinal micro flora, digestive disorders and even death in some cases. It was recorded that 2.2 million people killed from foodborne illness globally every year and the burden arising from foodborne diseases is larger (Kuchenmuller et al. 2009).
Antibioticsare massively used to overcome food poisoning; however, from health point of view they badly affect thenormal micro flora of gut but also microbes become antibiotic
resistance. The problem needs to be dealt with some other way like adding bio preservatives or antimicrobial agents in food.
To control microbes in foods, numerous methods have been adopted including the use of synthetic and natural antimicrobial agents. Scope of natural antimicrobial agents are increasing day by day and different natural sources are being utilized to get these agents. Among these natural sources milk is best and widely utilized source from long times. Milk contains many biologically active compounds among which lactoferrin is one of them.
Lactoferrin is a multifunctional globular glycoprotein from transferrin family, an iron-binding protein. It is part of innate immune system and has antibacterial activity known as far back as 1930. It was first isolated in 1939 from cow milk (Charrondiere et al. 2011). Lactoferrin belongs to the transferrin family having ability to bind iron two times higher than other transferrin proteins. Its molecular weight is 80 kDa and has about 700 amino acids depending upon species e.g. cow, buffalo, goat and sheep (Adlerova et al. 2008). Lactoferrin molecule consists of simple polypeptide chain folded into two symmetrical and highly homologous lobes (N and C) connected by a hinge region. Both lobes bind two metal ions in synergy with carbonate (CO32-). Not only Fe2+ and Fe3+ ions but Cu2+, Zn2+ and Mn2+ ions can also bind. Lactoferrin can bind Fe3+ reversibly so it can exist as free of Fe3+ (Apo-Lf) or in association with Fe3+ (Holo-Lf) and exhibits different three dimensional structure depending upon binding to Fe3+. Apo-Lf has an open structure and holo-Lf has closed which provide resistance to proteolysis. At iron-binding site Aspirin, two Tyrosine, and Histidine amino acids are directly involved in each lobe and Arginine is bound to CO32- ions. Number and position of Cystine-residues allows intermolecular disulfide bridges and Asparagine-residues in both lobes provide several sites for N-glycosylation (Farnaud and Evans 2003).
Lactoferrin is produced by mucosal epithelial mammary cells of human, cows, buffaloes, goat, horses, many other mammals and fish. It is widely distributed in body tissues and present in mucosal surfaces, specific granules of leukocytes and in biological fluids like tears, saliva, digestive fluids, seminal fluids and most abundant in milk comprising the second highest protein in human milk after casein. Concentration of lactoferrin in different species is for cow milk (80-500 mg/L), buffalo milk (50-320 mg/L), camel milk (200-728 mg/L), goat milk (98-150 mg/L) and sheep milk (20-140 mg/L) (Krol et al. 2011).
Many physiological functions of lactoferrin have been attributed. It plays an important role in iron regulation, non-specific immune response, regulation of cells growth and differentiation, protection from cancer, anti-inflammatory, anti-oxidant and strong antimicrobial activity against bacteria, fungi, yeast, viruses and parasites (Conneely et al. 2005). Another dominant role of lactoferrin is during involution of mammary gland. Concentration of lactoferrin increased dramatically from 0.1-0.3mg/ml in normal milk to 20-30mg/ml by 30 days in dry period. It is particularly important for bacteriostatic properties and non-specific defense against invading bacteria. Lactoferrin also affects phagocyte function and limit oxidative degeneration of cell components during inflammation and involution (Welty et al. 1976).
Lactoferrin exhibits strong antimicrobial activity against different bacteria, virus, protozoa, fungi and yeast (Hancock and Janssen 2009). The antibacterial activity of lactoferrin is due to two mechanisms; by binding the iron at infection sites, making it unavailable to bacteria and direct interaction of N-terminal of lactoferrin with micro-organism (Cruz et al. 2009, Orsi 2004). Lactoferrin acts differently with Gram-positive and Gram-negative bacteria (Sharma et al. 2013). It damages Gram-positive cell wall through interaction with negatively charged lipoteichoic acid causing reduction in negative charge on cell wall and favor contact between lysozyme and inner peptidoglycan (Fayad 2012). Gram-negative bacteria are destroyed by interaction of lactoferrin with external lipopolysaccharides by preventing contact with Ca2+ and Mg2+ ions which cause release of lipopolysaccharides, increase permeability and ensures damage (Ochoa and Cleary 2009, Ekins et al. 2004).
Milk and milk products are one of main diet in Pakistan and all over the world. During manufacturing different milk products, a number of by-products are obtained. Among them, cheese whey is produced in high volumes. It is commonly dumped off into sewerage which cause serious environmental problem as it contain high organic matter as well as loss of valuable nutrients it contain. Whey has Biological Oxygen Demand (BOD) ranges from 40,000 to 60,000 ppm (Sayadi et al. 2006) while permitted limit for BOD of domestic sewerage is 200 to 300 ppm. In order to overcome this problem there is need of effective and permanent way for treatment of whey. However, conversion of whey into non-food items like biogas is unreasonable as it is rich in unique nutrients.
Now-a-days there is an interest growing on to find new ways of whey utilization throughout the world. One option is to use the whey in processes in which saleable food or pharmaceutical products can be obtained. Whey could be subjected to different techniques to isolate different components like lactose, lysozyme and immunoglobulin. Likewise, lactoferrin can be isolated from cheese whey by cation exchange chromatography without loss of its biological properties in single step method and about 90% purity (Wu et al. 2011, Moradian et al. 2014).
In this study, we anticipated to use lactoferrin from bovine milk as natural antimicrobial agent. It has been shown that lactoferrin hasstrong antimicrobial activity against different bacteria, fungi, yeast, viruses and parasites (Conneely et al. 2005). In our country, very little work has been carried out onlactoferrin as natural anti-microbial agent. In fact, all over the world, the research scenario is now changing and concentrating toward the extraction of natural agents for product safety and health improvement. The lactoferrin has a potent anti-microbial activity against common foodborne pathogens. Due to the negative health effects of synthetic anti-microbial agents, the uses of natural sources are being encouraged all over the world. Our main focus of this study is to check the anti-microbial activity of lactoferrin against three pathogenic bacteria Escherichia coli, Staphylococcus aureus and Salmonella enteritidis isolatedduring our previous study.
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Production Of Single Cell Protein By Using Banana Peels As Substrate And Its Biological Evaluation In Broiler Chicks
Material type: Book ; Literary form:
Publisher: 2015 Dissertation note: The term single cell protein (SCP) refers to dead, dry microbial cells or total proteins extracted from pure microbial cell culture and is produced using a number of different microorganisms including bacterium, fungus and algae. It can also be called biomass, bioprotein or microbial protein.
Besides high protein content (about 60-82% of dry cell weight), SCP also contains fats, carbohydrates, nucleic acids, vitamins and minerals.
Fermentation media containing grinded banana peel as substrate was used to check the production of single cell protein for the selected Arachniotus sp. Different parameters were optimized for higher production of SCP e.g: Incubation period, pH, volume of inoculum, carbohydrate source, concentration of corn steep liquor and ionic salts concentration.
The biomass yield was estimated for total protein content by Lowrymethod. Biomass produced from fermentation was used for biological evaluation in feed trials of broiler chicks.
It is found that Arachniotus sp gave maximum single cell protein 7.49 g/L using 10 g banana peels at 72 hours incubation period. And protein concentration increased 7.58 g/L by optimizing volume of inoculum 2ml. It is observed in present study carbohydrate source also increases the protein concentration 8.41 g/L when carbohydrate source was optimized (glucose 3%).
Later on it was found that nitrogen source also enhance the protein production upto 12.61 g/L by using 2% corn steep liquor. Results also revealed that ionic salt concentration also play important role in the production of biomass protein, addition of 0.075% CaCl2.H2O produced 14.45 g/L single cell protein using above mentioned optimized conditions. 0.050 %
K2HPO4 produced 15.06 g/L. Addition of 0.050% MgSO4.7H2O produced maximum protein 15.86 g/L.
Biological evaluation in broiler chicks of this biomass protein shown there is no deleterious effects on weight gain, feed conversion ratio, protein efficiency ratio and net protein utilization. Maximum weight gain observed 215.6 grams in the group (C) in which 50% sunflower meal was replaced with biomass protein.
Feed conversion ratio in group (C) was 2.64 in which 50% sunflower meal was replaced by biomass protein and in group (B) was 2.51 in which 25% sunflower meal was replaced. And in control group (A) feed conversion ratio was 2.41.
Protein efficiency ratio was observed with non-significant value. And same results were shown by Chaves et al (1988) who reported non-significant differences among the standard and test diet when Chaetominumcellulolyticum biomass was fed to chicks. Net protein utilization observed in present study gave significant P value among the groups.
So it is concluded that single cell protein produced by this method is cheap and can be used in the food industry as food supplements and can also be included in poultry feed. The study findings suggested that microbial biomass produced by Arachniotus sp using banana peels as substrate can be replaced upto 50% of the protein supply by sunflower meal without any deleterious effects on growing broiler chicks. Moreover, it will also help in the reduction of pollution by using waste i.e. banana peel for useful purpose.
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Proteomic And Genomic Analysis Of Methicillin-Resistant Staphylococcus Aureus And Efficacy Of Indigenous Medicinal Plants Essential Oils
Material type: Book ; Literary form:
Publisher: 2015 Dissertation note: A Cohort study (prospective and observational) was performed to study the prevalence of Methicillin resistant Staphylococcus aureus from the healthy individuals of community, hospitalized patients and associated health-care workers and indigenous plants essential oils were screened as new, improved & potent antibacterial/s against resistant strains of MRSA.
The method involved isolation and identification of MRSA from surgical wounds of hospitalized patients & associated health care workers in a tertiary care hospital in Lahore and healthy volunteers from the community. Plant essentials oils & extracts were evaluated for their antibacterial activity against selected MRSA isolates. Oils were recovered by steam distillation using an all-glass distillation assembly. Then in vitro sensitivity and MICs of plant essential oils were determined using vancomycin and linezolid as commercial standards. The essential oils were screened further for the active constituents by column chromatography using various solvents and identification of compounds were performed by GC/MS analysis and the fractions which showed prompt results were evaluated for antimicrobial activity against the MRSA isolates in quest to find new therapeutic options. Finally effective essential oils and their active fractions were studied for their toxicity using in vitro Genotoxic assays such as Ames and Comet assays. To further ensure their beneficial effects antimutagenic effect of the essential oils were also studied.
Prevalence of S. aureus among patients was 52.9%, in HCWs 86.5% and in community 74% with an overall percentage of 72.6%. Among S. aureus those declared as MRSA were 91.8% from patients, 50.6% from HCWs and 59.5% from community with an overall percentage of 62.2% MRSA. Among the isolated MRSA overall 90.6% were Coagulase positive and 75.2% were biofilm positive.
The pattern of MRSA resistance against current antibiotics have shown an overall increase in the resistance with maximum shown for lincomycin followed by tetracycline, ampicillin, fusidic acid, amoxicillin and piperacillin with tazobactam. The most effective options among current regime were tigecyclin, amikacin and meropenem showing an overall least resistance. Resistance against linezolid was observed with an overall percentage of 25.6 % and vancomycin 33.3% by disc diffusion method.
The MRSA isolates resistant to one or more groups of antibiotics were declared as MDRs. Among patients and health-care workers all were declared as MDRs where as in community 93.1% isolates were MDRs.
Upon Protein profiling using whole cell proteins 44 bands of the polypeptides were produced with molecular size 10-200kDa from the three sampling groups and were categorized into 5 clusters showing an overall significance correlation with each other explaining an interesting fact that all these strains were interlinked establishing the fact of flow of hospital acquired MRSA in the community and vice versa. This analysis also gave an insight in explaining the fact of horizontal transmission of infection within the hospital.
Keeping in view the raise in resistance among current available antibiotics indigenous medicinal plants essential oils were screened for active constituents exhibiting anti-bacterial effects against MRSA isolates.
Maximum yield was obtained from Carum copticum followed by Cuminum cyminum and minimum yield was obtained in case of Zingiber officinale. Upon qualitative analysis of all five essential oils Carum copticum essential oil showed zones of inhibition greater than the standards vancomycin and linezolid followed Cuminum cyminum and Zingiber officinale in all three
sampling groups. Anethum sowa and Myristica fragrans essential oils showed no activity against MRSA.
Minimum inhibitory concentration of the three essential oils determined by micro broth dilution method indicated that Carum copticum showed least value in all three types of MRSA isolates followed by Zingiber officinale and Cuminum cyminum.
Effective essential oils were further fractioned using silica gel gravity columns. All the fractions obtained were screened for the anti-bacterial activity against all three types of MRSA isolates. Only fraction F1 of Carum copticum showed activity greater than pure essential oil and the two commercial standards of vancomycin and linezolid.
For the identification of active constituents GC/MS analysis was performed on all three essential oils and their respective fractions. In case of fraction F1 the most dominant constituents were Carvacrol, p-Cymene, Ʈ-Terpinene and Apiol. In other two plants none of the fractions were effective. Therefore it was concluded to use pure essential oils in case of Zingiber officinale and Cuminum cyminum rather than their individual fractions and incase of Carum copticum Fraction F1 has shown superior activity.
Finally these essential oils were tested for possible mutagenic effect using bacterial reversion mutation assay and Comet assay. No mutagenic effects were observed at MIC and above doses. These effective essential oils were also evaluated for possible antimutagenic effect. Both Carum copticum and Zingiber officinale essential oils showed strong antimutagenic effects and weak antimutagenic effect by Cuminum cyminum. Upon analysis of nuclear damage none of the plants essential oils and fraction F1 of Carum copticum showed genotoxic effects and indicated to be safe. Thus from the study it was concluded that Carum copticum essential oil and its fraction F1 were the most effective to be further investigated as an alternative treatment for MRSA infections.
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Sero-Screening Of Camels For Different Infectious Diseases
Material type: Book ; Literary form:
Publisher: 2015 Dissertation note: Camel is the precious and important animal in Pakistan. Camel is the most well adapted livestock species, survives and produces in climatic extremes and is well appreciated for its significance in the pastoral economy of the province. The camel being an important livestock species uniquely adapted to hot and arid environments and therefore contributes significantly to the food security of the nomadic pastoral households. Although camel being hardiest animal is less susceptible to diseases as compared to other livestock animals but literature shows that some diseases are still prevalent in camels. In view of the significance of camel as livestock animal as well as the symbol of cultural heritage of the nomadic pastoralists, there is a need to combat different diseases to which camels are susceptible and then appropriate control strategies should be applied.
Present study was designed to check the percentage positivity of different major diseases in camels that may pose serious issue relating to camel health and its importance as an important livestock animal. The diseases included in this study are Q fever, Brucellosis, FMD, CBPP and Neosporosis. ELISA is used to detect antibody prevalence by using specific kit based protocol for each disease whereas in case of Brucellosis RBPT is also used as basic screening test.
And it was found that Q fever has highest percentage seropositivity in both districts as compared to other diseases whose presence in camels was found to be almost seronegative.
So it was concluded that camel is still resistant to many diseases though some diseases are still prevalent in camels and these diseases should be controlled through public awareness and routine screening.
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Evaluation Of Microbiological Quality Ofshawarma Sold By The Street Vendors Of Lahore
Material type: Book ; Literary form:
Publisher: 2015 Dissertation note: Food safety is a scientific discipline describes handling, preparation and storage of food that prevents food borne diseases.It includes a number of ways that must be followed to avoid potentially severe health hazards.The tracks within this line of thought are safety between the market and consumer. With the fast pace of life the consumption of fast food is going to increase day by day in Lahore. It is imperative to prepare food at good hygienic conditions because of its perishability. The usual thougt is that food should be safe in the market and the concern is safe delivery and preparation of food for the consumer.
Microorganisms affecting food comes from natural micro flora or are introduced by manufacturing steps ranging from processing storage and distribution. In some cases these micro flora have no effect on the food and can be consumed without consequence, but those that are introduced during course of processing depending on type and level of contamination can spoil the food and cause food borne illnesses.
Food can transmit diseases from one person to another as well as serve as growth medium for bacteria that can cause food poisonoing. In the developed countries there are standards for food, whereas in less developed countries the main issue is the availability of safe water supply which is one of the critical item.
The present research work was undertaken to investigate the microbial count in shawarma which is a vended food. Samples collected from Anarkali,Wahdat road and Islampura showed high microbial count which shows contamination due to poor handling and hygienic conditions and improper storage conditionswhile the samples taken from Fortress, Gulberg having less microbial count as compared to these areas.
High microbial load in street foods occur due to improper food handling, unhygienic food preparation and processing, cooking and storage at inappropriate temperature. Consumer, food vendors and all type people should be aware of food hygiene, public health, implications of consuming contaminated foods, causative diseases. Food safety rules and implementation of food regulatory laws in food preparation, serving and preservation should be strongly maintained to avoid contamination problems and food-borne diseases.
Following points should be strongly observed and maintained for future work and further investigative study to improve the quality of shawarama.
1. Ensuring regular inspection and periodical check on these food preparation procedures.
2.Continuous lab test and analytical lab analysis to check the unwanted presence of any new
harmful agents in these foods to ensure food safety for consumers.
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Evaluation Of Microbial Quality Of Burger Sold By Roadside Fast Food Centers In Various Regions Of Lahore
Material type: Book ; Literary form:
Publisher: 2015 Dissertation note: Food safety actions need to cover the entire food chain, from production to consumption. The implementation of food safety include safety between industry and the market and then between the market and the consumer. In considering industry to market practices, food safety considerations include the origins of food including the practices relating to food labeling, food hygiene, food additives and pesticides residues, as well as policies for certification systems for foods.
Food industry regarding safety issue is one of the most unseen areas of policy in developing countries, especially in Pakistan. As a result, food systems in these countries are not always as well controlled and comprehensive as in the mechanized world. This situation is exacerbated by an ever increasing population, rapid urbanization and, most importantly, a lack of the economic and practical resources needed to hold a sound food safety system. As a result, people in developing countries are continuously facing a wide range of potential food safety risks.
Food borne illness is a disease caused by consuming contaminated food or drink.There are more than 250 known food borne diseases. Food can transmit disease from individual to individual as well as serve as a growth medium for bacteria that can cause food poisoning.
Today, people try to save as much time as possible due to the requisite to cope with many tasks within restricted time. In many cases people who have busy schedule don’t have enough time to cook food therefore prefer to eat fast food like burger, shawarma, fries etc. from fast food centers.
The present research was undertaken to investigate the microbial load of the chicken burger samples (n=96) collected aseptically from different regions of Lahore. Samples were collected in sterile plastic bags and analyzed for microbial load immediately after collection in Microbiology lab of University of Veterinary and Animal Sciences.
Nutrient agar was used to perform Total Plate Count, Mackonkey Agar was used for total coliform count, Manitol salt agar was used to count Staphylococcus Aureus and SalmonellaShigella agar was used to detect Salmonella.
The findings of the current study showed that samples taken multinational restaurant have less contamination than from local fast food centers. While among local fast food centers Liberty, Fortress, Anarkali and Akbarimandi have lesser microbial count than samples collected from Ichra, moon market, Mori gate. The difference in microbial load is due to improper food handling, processing, cooking and storage temperature, time and manufacturing practices of the food handlers.
After conducting my research I concluded that following points should be strongly observed and maintained to improve the quality of chicken burger in local fast food centers
• Ensuring regular inspection and periodical check on these food centers
• Continuous lab test and analytical lab analysis to check the unwanted presence of any new harmful agents in these foods to ensure food safety for consumer.
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Antibiotic Resistance Pattern Of Staphlococcus Aureus And Its Resistance Modulation Using Medicinal Plant Extracts
Material type: Book ; Literary form:
Publisher: 2016 Dissertation note: This project was designed to evaluate the antimicrobial efficacy of Chloroform and
ethanol extracts of Calotropis procera (C .procera) and Eucalyptus globulus (E. globulus)
against Multiple Drug Resistant (MDR) Staphylococcus aureus isolated from human origin. This
study was conducted to evaluate the antimicrobial potential of C. procera and E. globulus alone
and in combination with antibiotics to check synergism between medicinal plants and resistant
S.aureus is a major pathogen which causes various infections. Infectious diseases affect
millions of people around the world and in the history these diseases are major cause of mortality
and morbidity across the globe. In past few decades rate of mortalities are continuously
increasing because of acquired resistance of S. aureus against multiple drugs, thus it is utter need
of time to discover some alternatives to antibiotics so that we can resolve this dilemma of
antibiotic resistance. Plant extracts are hope for this purpose as they have many compounds
which have potential to lower down the number of micro-organisms. Plants have benefits over
other as they are non toxic, non-reactive and have least side effects.
Total 20 samples of human origins were procured from Department of Microbiology,
UVAS Lahore and were subjected to check their antibiotic resistance profile against
Erythromycin, Amoxicillin and Ciprofloxacin by Kirby Bauer disc diffusion assay. Out of 20,
nine resistant isolates were separated. Among them three were resistant to Erythromycin, three to
Amoxicillin and three to Ciprofloxacin.
Leaves of C. procera and E. globulus were processed in Chloroform and ethanol
Solvents. Antimicrobial activity was evaluated by agar gel well diffusion assay in which zone of
inhibitions were measured. Minimum inhibitory concentration (MIC) of plant extracts was
evaluated by micro broth dilution method. Best antimicrobial activity was observed by ethanolic
extract of E. globulus.
Then combine effect of sub-inhibitory concentrations (SICs) of plant extracts and
minimum inhibitory concentration of antibiotics were determined by Well Diffusion assay. Four
different sub inhibitory concentrations of plant extracts i.e.10μg/ml, 20μg/ml, 40μg/ml and
80μg/ml were used in combination with fixed concentration of antibiotics i.e.100μg/ml to check
combinational effect of both. At selected sub-inhibitory concentration plant extract alone did not
show any antibacterial activity. Two of the isolates had shown modulation when amoxicillin and
plant extracts combination was used against them. The isolate labeled as S.aureus 4 showed
modulation with the use of Ethanolic extract of Calotropis procera and S.aureus 5 had shown
modulation with the use of chloroform extract of Calotropis procera. For further confirmation
two more concentrations of 160μg/ml and 320μg/ml were used along with100μg/ml Amoxicillin
against same isolates S.aureus 4 and 5. Zone of inhibition was observed with increased diameter
indicating modulation of two isolates. While erythromycin and ciprofloxacin resistant isolates
didn’t show any modulation.
Data of antibiotic resistance and resistance modulation using plant extracts was analyzed
by one way analysis of variance (ANOVA) followed by Duncan’s multiple range(DMR)
posthoc test using Statistical package for social sciences (SPSS) 17.0 Statistical software at α <
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Amelioration Of Pathological Effects Of New Castle Disease By Aloe Vera
Material type: Book ; Literary form:
Publisher: 2016 Dissertation note: Poultry industry has many threats from the infectious diseases. Newcastle disease is the most important disease of the poultry as it is distributed all over the world and it can cause huge economic losses in poultry industry. It is caused by the Newcastle disease virus (NDV) that can infect about 240 species of birds. Newcastle disease cause immune suppression in birds. It is reported that supplementations of Aloe vera enhances the immune status and reduce inflammation. So this research project was design to observe the effect of Aloe vera on lymphoid organs, growth performance and antibody response in Newcastle disease challenge birds. For this research a total of 120 broiler chicks were divided into four groups A, B, C, and D. Group A was control group while B and C were treated with 2 percent aqueous extract of Aloe vera. Group C was also vaccinated against New Castle disease. Aloe vera was given to group B and C from day one to end of study trial. Both of the groups were challenged with ND virus at day 21. Group D was vaccinated against ND and was challenged with ND virus at day 21 without supplementation of Aloe vera. Blood samples were collected at day 1, 7, 20, 24, 26 and 28 to determine the antibody titer against ND. Highest antibody titers were observed in group C as compared to all other groups which was vaccinated against ND along with supplemented with Aloe vera. For gross pathology and histopathology, lymphoid organs were collected at day 24, 26 and 28..The average feed intake of group A and D was significantly higher than group B and C before challenge of virus but the body weight gain of 2% Aloe vera supplemented broiler was significantly (p<.05) higher than without treatment of Aloe vera. The FCR of birds supplemented with Aloe vera treatment was significantly different from the birds without Aloe vera treatment. The FCR value of group C and B was higher than A and D.
A significant difference was observed in the weight of lymphoid organs of birds treated with Aloe vera as weight of organs was less in group C followed by group B, group A and group D. This was due to anti-inflammatory effects of Aloe vera. Microscopic examination revealed congestion, depletion of lymphocytes, dysplasia of thymic lobules, thinning of cortex, focal necrosis, disappearance of lymph follicles and inter-follicular edema like lesions within lymphoid organ of the groups challenged with Newcastle disease virus. However, cellular hypertrophy and decreased lymphocytes population were prominent changes in lymphoid organs of broiler treated with 2% Aloe vera. To check the virucidal effects of Aloe vera, a separate experiment was conducted in which 9 day old embryonated eggs were inoculated with ND virus along with 2 percent Aloe vera gel extract after incubation at 37Cº for an hour in group A while only ND virus was inoculated in group B. Candling was performed to see the survival of embryos in both groups which revealed a significant difference i.e. 16 percent embryos were found dead in group A while 80 percent was found dead in group B. Later on the amnioallantoic fluid of the eggs was used for spot Haemagglutination test. Group A showed less agglutination activity then group B. From this study it was concluded that Newcastle disease caused immune suppression and damage of vital organs in broiler while Aloe vera have immunomodulatory, anti-inflammatory and antiviral effects as it raised antibody titer against Newcastle disease virus and lowered the inflammatory processes along with inactivation of ND virus. It also promotes growth performance of broilers and helps the birds to survive against lethal ND disease.
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Pathogenesis Of Field Isolates Of Mannheimia Hemolytica In Experimentally Infected Rabbits
Material type: Book ; Literary form:
Publisher: 2016 Dissertation note: Shipping fever is one of the most economically important infectious diseases of ruminants with a wide prevalence throughout the continents. The disease is characterized by an acute febrile course with severe fibrinous bronchopneumonia. Infected animals may die within a few days of the onset of clinical signs, but those which survive the acute attack may become chronically infected. Both Mannheimia and Pasteurella species are commensally resident in the respiratory tract of healthy ruminants and are capable of causing infection in animals with compromised pulmonary defense system. Bovine respiratory disease (BRD) is the most common and costly problem encountered in stocker or feedlot calves. BRD also called “shipping fever”, accounts for major economic losses to the producer by reducing average daily gain, feed efficiency, and overall performance of beef calves. The aim of present study was isolation of M.haemolytica from cattle. The identification of organism was performed through biochemical tests and confirmation by polymerase chain reaction. The nature of disease was evaluated through gross and microscopic lesions.
A total of 50 tissue samples (25 lungs and 25 pharynx) were collected from Punjab Agriculture and Meat company Lahore and brought to the Department of Pathology UVAS, Lahore and were analyzed for biochemical and molecular detection of M .haemolytica. For studying the pathogenesis of the disease, experimental infection was given to rabbits in Department of Pathology, UVAS Lahore. Rabbits were randomly divided into Group A, Group B and Group C with nine rabbits (n=9) in each group. Experimental infection of field isolated M. hemolytica was given intratrachealy to the rabbits. Rabbits of group A and B were infected with 0.5 mL bacterial inoculum having 103 and 106 CFU/mL respectively. The rabbits of Group C served as control group. Rectal temperature of each rabbit was recorded daily. On postmortem,
gross and microscopic lesions were recorded.
The results showed that rabbits of control group not showed any gross or microscopic change. There was significant increase in rectal temperature of infected rabbits as compared to uninfected rabbits. The gross lesions were specific for the organism which was prominently observed in lungs of rabbits. The microscopic lesions revealed that there was severe consolidation, congestion and fibrin exudation in lungs of rabbits of group A which were given less number of organism and they developed clear signs of disease. The rabbits of Group B showed less prominent signs compared to group A due to early death of rabbits. There were multiple hemorrhages, of varying sizes and hyalinization of myocardial cells in infected rabbits. The severity of changes was significantly more different in Group A, as compared to Group B.
It can be deduced by this study that the rabbit can be used as a model for further studies exploring the pathogenesis of the disease as the lesions resemble to shipping fever caused by M. hemolytica in ruminants. The lesions, which developed, could be descending infection resulting in typical lesions of bronchopneumonia or lobular pneumonia.
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Antibacterial Activity Of Plant Extracts Against Antibiotic Resistant Pseudomonas Aeruginosa And Their Cytotoxicity Profile
Material type: Book ; Format:
; Literary form:
Publisher: 2016 Dissertation note: Pseudomonas aeruginosa is a common opportunistic pathogen of respiratory tract
and cause both hospital and community acquired infections. For the treatment of
infections antibiotics are used but due to random selection of commonly used antibiotics,
resistance in P. aeruginosa has developed. This problem may leads to the high morbidity
and mortality rate. Different medicinal plants have antibacterial activity in their
secondary metabolite. Secondary metabolites are terpens, flavonoids, alkaloids and
alcohols etc. So the plant extract could be the alternative therapy for the treatment to
reduce the antibiotic resistance problem.
Isolates of P. aeruginosa was procured from the main clinical laboratory of Mayo
Hospital, Lahore and identified biochemically according to bergey’s manual of
determinative bacteriology. Antibiotic resistance pattern of identified P. aeruginosa was
evaluated by Kirby Bauer disc diffusion assay against selected antibiotics includes
ciprofloxacin, levofloxacin, meropenem and imipenem. Measure the zone of inhibition
and isolates marked as resistant, intermediate and sensitive. Resistant strains were
alienated for further evaluation.
Leaves of Eucalyptus globulus (Tasmanian blue gum) and Calotropis procera
(apple of Sodom) proceed for extraction and the plant extracts was obtained by using
solvent chloroform and ethanol. Percentage yield of both plant extract was calculated.
High percentage yield was obtained from Eucalyptus globulus and less percentage yield
was gained from Calotropis procera in comparison The obtained extract was dried and
the resultant material was used in well diffusion assay to evaluate the antibiotic
sensitivity of resistant P. aeruginosa against selected plants. Stock of plant extracts was
prepared by dissolving 1g of plant extract in 1ml of DMSO. Well diffusion assay was
performed and zones were measured in millimeter and categorized as resistant, sensitive
and intermediate. Isolates that are susceptible to plant extracts were separated and
Minimum inhibitory concentration of susceptible isolates was determined by broth micro
dilution assay and cytotoxicity profiling was done by 3-(4, 5-dimethyl-2-thiazolyl)-2,5-
diphenyl-2H-tetrazolium bromide (MTT) assay and cell survival percentage was
Data recorded during the study was analyzed by one-way analysis of variance
(ANOVA) followed by Duncan’s test using the SPSS statistical software program.
Differences were considered significant at P < 0.05.
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Evaluation Of The Microbiological Quality Of Ice Cream Sold At Local Shops In Lahore
Material type: Book ; Literary form:
Publisher: 2016 Dissertation note: Ice cream is a delicious, wholesome, nutritious frozen dairy food. It is noticed that generally manufacturing of ice cream on small scale production units like local shops doesn't totally follow the standard procedures of ice cream production. Ice cream can be contaminated with microorganisms if some ingredients have been added after pasteurization or by means of improper sanitation of the equipment and environment. Microbiological quality of ice cream reflects hygienic practice in production and is an indication of food safety. There was variation in quality of locally produced ice cream in different areas of Lahore.
Food safety is a scientific discipline describes handling, preparation and storage of food that prevents food borne diseases. It includes a number of ways that must be followed to avoid potentially severe health hazards. The tracks within this line of thought are safety between the market and consumer with the fast pace of life the consumption of fast food is going to increase day by day in Lahore. It is imperative to prepare food at good hygienic conditions because of its perishability. The usual thought is that food should be safe in the market and the concern is safe delivery and preparation of food for the consumer.
Microorganisms affecting food comes from natural microflora or are introduced by manufacturing steps ranging from processing storage and distribution. In some cases these micro flora have no effect on the food and can be consumed without consequence, but those that are introduced during course of processing depending on type and level of contamination can spoil the food and cause food borne illnesses.
Food can transmit diseases from one person to another as well as serve as growth medium for bacteria that can cause food poisoning. In the developed countries there are standards for food, whereas in less developed countries the main issue is the availability of safe water supply which is one of the critical item. Foodborne illness is a problem resulting from the consumption of contaminated food, bacteria, viruses, or parasites, natural toxins, chemicals that contaminate food.
For this study total 108 locally produced samples of ice cream were carried from different areas of Lahore. Mughalpura, Sadar, Model town, Gulberg, Town ship and Johar town. 6 shops from each area and 3 samples from each shop were purchased and put in the sampling box with ice packs. Samples were analyzes for microbial load in microbiology Lab of University of Veterinary and Animal Sciences.
Each sample was analyzed for microbiological analysis (Enumeration of Total Viable Count (TVC), Enumeration of Total Coliform Count (TCC), Enumeration of Total Staphylococcal Count (TSC) and detection of salmonella spp.
Nutrient agar was used to perform Total plate count, Mackonky agar was used for Total coliform count, Mannitol salt agar was used to count S.aureus , Salmonella Shigella agar for Salmonella detection . Each analysis was performed 3 times.
The samples collected from Mughalpura, Sadar and Model town were highly contaminated having more microbial count which shows contamination due to poor handling and hygienic conditions while Gulberg, Johar town and Town ship having less contaminated.
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Comparative Quality Evaluation Of Raw And Pasteurized Milk
Material type: Book ; Literary form:
Publisher: 2016 Dissertation note: This particular project was designed to evaluate the overall quality of raw and pasteurized milk available at different areas of Lahore. The parameter which was checked includes microbiological analysis, adulterants, physicochemical properties and the effect of temperature on vitamin C in milk samples. Raw samples were collected from ten different towns of Lahore, whereas pasteurized milk samples belong to ten different brands. Ten samples were collected under control conditions from animals in sterilized containers.
For microbiological analysis four parameters were selected including total plate count (TPC), total coliform count (TCC), total psychrotrophic count (TPSC) and total yeast and mold count (TYMC) whereas, different adulterants like adulteration test was done such as urea, starch, hydrogen peroxide, detergent or soap, sorbitol, quaternary ammonium compound, boric acid, cane sugar, sodium chloride, formalin and hypochlorite were checked by using the milk adulteration kit in QOL. Milk contains casein and whey proteins. Whey protein being added in the milk to increase its density which is considers being an adulterant. In this project whey protein was estimated in all milk samples by titration method. Physicochemical characteristics of milk are an important parameter to judge the quality of milk. These physicochemical properties include fat%, SNF%, density kg/m3, lactose%, solid/ash, protein% and pH. Physicochemical properties were evaluated mechanically by Milkoscan. Heat treatment is an important method to reduce the microbiological contamination of milk. These treatments may include pasteurization and UHT etc. During the heat treatment some of the micronutrients may deteriorate thus compromising the quality of milk. Vitamin C is among those heat labile micronutrients. Vitamin C was checked quantitatively in market and self-collected samples by using titration method.
It was concluded that total plate count TPC, TCC, TPSC and TYMC of raw milk samples were above the standard value indicating the poor quality of the milk. As far as the pasteurized milk samples were concerned ninety percent of the samples showing higher values for TCC, TPSC and TYMC. Total plate counts of all self-collected raw milk from a healthy animal were found within the standard value. Counts were in range of 3.8x 103 – 8.9x103 CFU/mL of all milk samples. TPC of all self-collected raw milk from a healthy animal were found within the standard TCC were found within permissible value (102 CFU/mL .TPSC were negative for all milk samples. TYMC were in range of 2.6x101 -7.2x101 CFU/mL. Among milk samples (n=10), three samples (30%) were positive for TYMC were while remaining samples (70%) were negative and showed no growth.
Physicochemical factor show that 50 percent of raw milk have low nutritional value as compared to the standards which are buffalo and cow milk contains 7.6, 4.5% fat, 3.8, 3.8 % protein, 5.1, 4.9% lactose, 0.78, 0.72% ash and 17.0, 13.9% total solid respectively. In raw milk mean of fat (%), solid not fat (%), lactose (%), Solid/ash (0%), protein(%) and pH were 4.50±0.03, 7.915±0.06, 23.05±0.055, 3.893±0.06, 3.85±0.05, and6.9±0.0.02 respectively. In pasteurized milk mean value for fat, SNF, lactose, ash, protein and pH were 3.48 ±0.13, 7.24±0.10, 3.60±0.05,0.5 ±0.06, 2.82±0.05, 7.2±0.20 respectively. Pasteurized milk is good for consumption.
Different adulterant such as urea, starch, hydrogen peroxide, Sorbitol, QAC, Boric acid, Cane sugar, NaCl, Carbonate, Formalin, hypochlorite, whey protein, Added water and soap /detergents were evaluated in all milk samples. Among these adulterant water (66%) was found in majority of milk samples, followed by whey protein (15%), starch (13%), (10%) NaCl and
(8%) cane sugar were detected in raw milk samples. n Pasteurized milk samples only added water (49%) and whey protein (31%) was detected.
Among the raw milk samples the maximum and minimum concentration of vitamin C was observed 0.33±0.02 and 3.33 ±0.02 mg/100ml and for pasteurized milk maximum and minimum concentration of vitamin C was observed 2.54mg/100ml and 0.32 ±0.02 mg/100ml respectively. In self- collected samples the minimum and maximum concentration of vitamin C was observed 5.25±0.02 and 8.34 ±0.04 mg/100ml respectively and after pasteurization in laboratory minimum and maximum concentration of vitamin C was observed 3.48±0.04 and 5.83 ±0.02 mg/100ml respectively. These observations had showed that pasteurization treatment decreased Vitamin C quantity.
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Isolation, Molecular Identification And Antibiotic Resistance Pattern Of Salmonella Enterica From Fancy Birds
Material type: Book ; Literary form:
Publisher: 2016 Dissertation note: Salmonellosis is a disease with serious health issues related to food borne illness and
most of world’s population is suffering from it. Early diagnosis in case is very important for
treatment of disease. Salmonellosis may hidden as a carrier state, acts as zoonotic components
for transmission of disease. Therefore the test with more diagnostic value needs to be developed
like Polymerase chain reaction after culturing and microbiological examination.Salmonella
enterica infections continue to pose a significant risk for poultry industry and fancy birds.
Salmonella infections have been controlled by antibiotics but in recent times antibiotic resistance
in microorganisms especially in Salmonella is a global health issue. Antibiotic resistant
Salmonella has further compounded the problem. Poultry isolate of Salmonella enterica (n=150)
were procured from Jallo park, Safari park and household pets which are taken to Pet Centre
University of Veterinary and Animal Sciences Lahore then brought to Department of
microbiology UVAS Lahore and identified by biochemical testing, morphology, staining
characters and genus specific PCR. Antibiotic Susceptibility was checked by disc diffusion
method against amoxicillin (30μg), ampicillin (10μg), cefixime (5μg), , ceftazidime (30μg),
ceftriaxone (30μg), ciprofloxacin (5μg), gentamicin (10μg), nalidixic acid and tetracycline
(30μg) and resistant pattern was 100 % in ampicillin and tetracycline and 41.18% and 58.82% %
in gentamicin and ciprofloxacin respectively while antibiotic show 0% resistance. Fancy birds
are carriers of drug resistant Salmonellae.
A total of 150 samples collected from Zoo Lahore, safari park and household pet fancy
birds each of n=50. Samples will enriched by non-selective and selective media, After isolation
on selective media macroscopic, biochemical analysis and microscopic examination done. DNA
extracted from culture isolated from cloacal swabs and polymerase chain reaction performed
using primers. Amplication will be observed using Agarose gel electrophoresis.
Research highlighted the prevalence of Salmonella in fancy birds and its possibility of
transmission to human beings. Research also provided data on antibiotic resistance in
Salmonellae from fancy birds and its possible role in ever increasing problem of antibiotic
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Pathogenesis Of Aflatoxin B1 In Quails Under Experimental Conditions And Detoxification By Biological And Chemical Means
Material type: Book ; Literary form:
Publisher: 2016 Dissertation note: Secondary metabolites of certain fungi produce toxins under favorable conditions especially while growing on different food grains. Mycotoxins are among major threats to growing poultry industry and human beings. Aflatoxins are closely related, biologically active fungal metabolites and commonly produced by Aspergillus species.
A research was carried out to evaluate the ability of Aspergillus flavus for Aflatoxin B1 production using rice, wheat and maize as substrates. Lethal effects on growth performance parameters, hematological and histopathological of graded doses of aflatoxin B1 in quails under experimental conditions were observed. Effect of Aflatoxin B1 on humoral immune response to Newcastle Disease virus vaccine in quails were determined. Biological detoxification of Aflatoxin B1 by Saccharomyces servisiae was evaluated in quails. Comparative evaluations of different commercially available toxin binders were checked. All these experiments were carried out till the six weeks (42 days).
Aspergillus flavus was identified on the basis of macroscopic and microscopic characteristics. Rice, wheat and maize grains was used as substrate to check the level of Aflatoxin B1 produced by inoculating an aqueous suspension of 106 spores/ml. Aflatoxin B1 checked by Thin Layer Chromatography (TLC) and quantified by High Performance Liquid Chromatography (HPLC).
Quails were reared under standard management conditions in five groups (A, B, C, D and E) having sixty each. Each group was further divided in two independent units. Diets offered to groups were control (without toxins), 0.25, 0.50, 1 and 2 mg Aflatoxin B1/kg feed. One unit of
each group was vaccinated with Newcastle Disease Virus (NDV) vaccine while other was not and studied the lethal effects on growth performance, blood parameters, immune response and histopathology of vital organs. At the end of the experiment, it was found that the deleterious effects of Aflatoxin B1 were dose and duration dependent. As the level of the toxin was increased, the lethal effects were prominent. The growth performance parameters including gain in body weight, feed intake and feed conversion ratio was adversely affected at high doses. The body weight gain was significantly reduced in Aflatoxin B1 treated groups as compared to control group. Similarly feed intake and feed conversion ratio were significantly different from the control group. The hematological studies exhibited that aflatoxin B1 significantly reduced the hemoglobin, packed cell volume and total leukocyte count whereas the erythrocyte sedimentation rate was significantly increased as compared to control group. The immune response against NDV vaccine was adversely effected in Aflatoxin B1 treated groups and values of Antibody titer in AFB1 were significantly low as compared to group A( control) In the second experiment, Saccharomyces cervisae (SC) dried powder was mixed in basal quail diet having 0.5mg Aflatoxin B1 for all experimental groups and control was without toxins. SC was added at levels of 0.5 gm, 1.0 gm and 2.0 gm /kg of feed. It was recorded that Saccharomyces cervisae (yeast) have the potential to remove the deleterious effects of Aflatoxin B1. Yeast effectively detoxified the Aflatoxin B1. The results recorded of growth performance and other parameters were non-significantly different from the control group. Chemical detoxification of Aflatoxin B1 was evaluated in quails using commercially available toxin binders. Toxin binders used were activated charcoal, kaoline, Myco AD and selenium plus vitamin E and mixed in basal quail diet having 0.5mg Aflatoxin B1 for all experimental groups and control was without toxins. The Myco AD and selenium plus vitamin E showed the highest detoxification potential as compared
to other chemical toxin binders. Groups E and F showed the results of growth performance, hematological, immune response and histopathological were non-significantly different from the control group (A). Kaolin was moderately detoxifying the toxin.
Presence of aflatoxin B1 in soft tissues was checked by TLC and quantified using HPLC. The liver exhibited the residues of Aflatoxin B1 at high doses of toxin. Group D and E rearing on feeds having 1mg AFB1 /Kg feed and 2mg AFB1 /Kg feed of toxin showed the residues of AFB1 in liver and kidney.
Statistical means for growth performance parameters, hematological, immune response and histopathological scores in each subunit of quails were analyzed by applying one way ANOVA and Duncans‟s Multiple Range (DMR) test at 95% probability. Aflatoxin B1 is lethal and lowers the performance of birds. The lethal effects can be detoxified by biological and chemical means to lower the economic losses to poultry industry. It can be concluded that biological detoxification is preferably better as compared to chemical detoxification.
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Identification And Molecular Characterization Of Mycobacterium Bovis And Mycobacterium Paratuberculosis In Wild Cats.
Material type: Book ; Literary form:
Publisher: 2017 Dissertation note: Cases of wildlife diseased with Mycobacterium species are existing in Pakistan and result in high morbidity and mortality. Vaccine is the only preventive measure, but in wildlife the vaccine administration is a strenuous job. In Pakistan vaccination practice is not up to the mark and vaccination schedules are not being followed. Mycobacterial diseases have gain popularity due to their zoonotic effect. Scat samples from Lahore Safari Zoo and Lahore Zoo were collected and properly labelled. Conventional PCR along with Touchdown PCR was done using universal primer sets of M. bovis and M. paratuberculosis. The amplicons were run on agarose gel and the bands were observed under Gel Doc system.
The objective of the study was to detect the currently prevailing Mycobacterium bovis and mycobacterium avium subspecies paratuberculosis in wild cats in Pakistan.
However the results obtained from different kinds of PCR were negative, showing that the wild cat population of Lahore Zoo Safari as well as Lahore Zoo were free from Mycobacterium bovis and mycobacterium avium subspecies paratuberculosis.
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Molecularcharacterization And Antibiotic Resistance Pattern Of Pathogenic Eschericia Coli From Fancy Birds
Material type: Book ; Format:
; Literary form:
Publisher: 2017 Dissertation note: Colibacilosis is a disease which is really harmful as food borne illness and many population of earth has been suffering from it. If we do early diagnosis then it would be easy to identify the disease. E. coli is an opportunistic bacteria and it can easily cause disease in favorable conditions and may also act as zoonotic issue. So the persons having birds as pets in their houses should consider for the diagnosis. So, for this purpose the more accurate tests like polymerase chain reaction should be present. E. coli can cause serious risks for poultry industry and fancy birds.
Colibacillosis is caused by Escherichia coli with serious threats related to food borne illness and most of world’s population is suffering from it. Fancy birds are kept in homes as pets, zoo and safari parks for entertainment of visitors. Fancy birds may be source of transmission of pathogenic E. coli to human beings.
150 cloacal samples were taken and the process of isolation, identification, antibiotic resistance pattern and molecular characterization was done for pathogenic E. coli.
150 samples from which 50 samples were from Pet Center University of Veterinary and Animal Sciences, Lahore, 50 samples were from Lahore Jalo Park and 50 samples were from Safari Park Lahore. Samples were shifted to enrichment media and then selective and differential media. After isolation microscopic characters was observed and biochemical identification was performed.
Pathogenic isolates were collected and antibiotic resistance pattern was observed. After this DNA extraction was done, extracted DNA was observed on gel electrophoresis and then PCR was performed for toxigenic genes of E. coli.
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Pathological Effects Of Natural And Experimental Lead (Pb) Toxicity In Lohi Sheep At Jhang, Pakistan
Material type: Book ; Literary form:
Publisher: 2017 Dissertation note: Heavy metal toxicity is increasing due to increasing trends of urbanization and
industrialization. Lead poisoning has been recognized as a major public health risk,
particularly in developing countries. It is nephrotoxic, hepatotoxic, neurotoxic, carcinogenic
and mutagenic for animals and human. Sewerage water, fertilizers, leaded-gasoline and lead
based batteries are the sources of lead contamination in soil and forage. The lead particles are
taken up by animals from contaminated forages and excreted in animal products like milk and
The presence of Pb in drinking water, waste water, plant products and animal
products has been studied which is a serious risk for animal and public health. The
legislations for the disposal of household wastes and industrial effluents are very poor in
Pakistan. The calculation of safe Pb levels in different products is still to be needed.
Pathological effects of higher Pb levels have not been studied in Pakistan. The present study
was aimed to unveil the toxic effects at constant dose of Pb over a period of three months in a
local sheep breed of Pakistan. The status of Pb toxicity was also investigated in a polluted
area around sewage drain and mutton slaughter house at District Jhang, Pakistan.
The Pb concentration in soil, forage and irrigating water was found to be below the
permissible limits and was safe for agriculture but long-term ingestion of low Pb
concentration may have cumulative effect. The serum Pb concentration was found to be
above the recommended safe limits for producing Pb toxicity in animals. The different tissues
like kidney, liver and skeletal muscles also contained higher Pb level from the permissible
limits and found to be unsafe for public use. Kidney showed the highest Pb concentration and
the muscle contained the least Pb level in the present study.
The erythrocyte count, hemoglobin concentration and packed cell volume showed
inverse correlation with Pb concentration and mean values were below the normal range in
Pb treated sheep but anemia was not developed. The erythrocyte sedimentation rate was also
influenced by given dose of lead acetate during third month of treatment. The white blood
cells also revealed no effect on given dose of lead acetate in Lohi sheep in this study.
The biochemical parameters of field and treatment group showed higher concentration
as compared to control group of Lohi sheep but their means were falling within the normal
range of reference values. The disturbed biochemical parameters in apparently healthy sheep
with higher serum Pb concentration were indicative for liver and kidney damage.
Lohi sheep exhibited less effect on given dose of lead acetate during first two months
but more pronounced changes of chronic Pb toxicity were observed during last month of trial.
The histological changes were not observed on early period in lead acetated treated sheep.
The characteristic histological changes were observed on last slaughtering at day 90 in kidney
and liver including degeneration and focal areas of necrosis, dilatation of blood vessels with
accumulation of red blood cells and fibrosis in some areas. The nuclear changes were more
typical with intranuclear inclusion bodies in renal tubular epithelial cells but less
distinguishable in hepatocytes.
It was concluded that soil, forage and water contained low Pb levels in the study area.
The ingestion of low Pb level for longer period had cumulative effect in animals. The animals
might be resistant to low Pb level but their products are a severe risk for public health. So the
necessary measures should be adopted to minimize the heavy metal contamination in animal
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Molecular Characterization of Carbapenem Resistant Pseudomonas Aeruginosa Isolated from a Tertiary Case Hospital in Lahore
Material type: Book ; Literary form:
Publisher: 2017 Dissertation note: Empty CD.
Availability: Items available for loan: UVAS Library [ Call number: 2834-T] (1).
Assessment Of Toothpaste From Local And Imported Origins For Its Heavy Metals And Minerals Composition
Material type: Book Publisher: 2017 Dissertation note: Toothpastes are molten form substance used by toothbrushes to clean our teeth for personal hygiene. Toothpaste is very essential for our daily hygienic routine. Heavy metals can be found in toothpastes by different means like usage of raw materials for manufacturing in which they are present due to their persistence nature, plant and water source and deliberate addition. Their long time exposure can cause serious issues in humans. Above than permitted level of heavy metals and minerals cause many health problems. They can disrupt living systems that can lead up to cancer and death threat. High levels of concentration of heavy metals are not acceptable in toothpastes and many countries including Pakistan have proposed their limits that should not be surpassed.
The purpose of this study was to assess the concentrations of heavy metals and minerals in different brands of toothpaste to be safe for human use. Samples were collected from different markets and pharmacies of Lahore. Samples were divided into five groups; local regular, local herbal, imported regular, imported herbal and medicated. Each group has five toothpastes. Samples were digested by diacid method. One gram sample was taken in Kjeldahl’s digestion tube and 7mL of HNO3 was added and after 30 min 3ml of HCLO4 was added. This solution was heated at 150 °C for 30 minutes on digestion block and then at 250 °C until wine green or water clear color end point. Digested samples were analyzed by using Multi-channel Flame photometer (AFP 100) for Na, K and Ca. Heavy metals like Pb, Cu, Zn, Cr, Mn Ni, Se and Fe byAtomic Absorption Spectrophotometer (Polarized Zeeman Z-8230 Atomic Absorption Spectrophotometer).Flouride was determined by spectrophotometer (V-1100 Spectrophotometer). The result showed that lead (Pb) was under the permissible limit by Pakistani standard in all toothpastes. Copper (Cu) was not detected in any of the toothpaste. Zinc (Zn)concentration range was 0-137.9 ppm. In almost half of the samples zinc was not present at all while all other toothpastes have higher concentration than permissible limits. Forhan’s has the highest concentration (137.9 ppm) than permissible limit of US Patent.Although,mean value of every group has surpassed the permissible limit.Chromium (Cr) was also present above than permitted level in all of the toothpastes ranging 4-17.3 ppm. Manganese (Mn) concentration range was 0-37.5 ppm. Manganese was not found in 15 samples but in all others it has surpassed the permissible limit by Nigerian standard. Shield peppermint has the highest concentration of Mn. Mean value of every group has surpassed the permissible limit.Nickle (Ni) was present above than permissible limit in all of the toothpaste set by Nigerian standard. Ni concentration range was 1.6-10.8 ppm. Himalaya mint fresh has highest concentration of Ni. Selenium (Se) was present between 0- 1.5 ppm. Selenium (Se) was not found in six samples at all and three samples were having permissible limits which are; medicam herbal, himlaya mint fresh and sensdyne. While in all other samples it has surpassed the limit set by Chinese standard.Shield peppermint has highest concentration (1.5 ppm) of Se. Mean value for four group has surpassed the permissble limit while one group (imported herbal) has under the permissible limit.Iron (Fe) has passed permitted limit set by Nigerian standards in all of the toothpastes. The concertation range for iron was between 2.8-10.9 ppm.Forhan’s has the highest concentration. Mean value of every group has surpassed the permissible limit. Sodium (Na) has concentration range between 1425- 22608 ppm.Sodium (Na) was present above than permissible limit (2000 ppm) by US patentmedicam having the highest concentration except close up deep action, doctor herbal, oral b, dabur red and Himalaya sensi-relief that have under permissible limit but mean values of four groups have surpassed the permissible limit while imported herbal was present under the permitted limit. Potassium (K) was present between 145.8- 15283.3 ppm range. All of the toothpastes have under the permissible limit level of K. Calcium (Ca) was found in permitted limit in nine samples while all other have concentrations above than permissible limit(1120 ppm)by European standardmedicam having the highest concentration. Mean value of every group has surpassed the permissible limit. Fluoride was found in permissible limit in 15 samples while in all others it has surpassed permissible limit (1500 ppm) set by Pakistani Standardsensodyne having the highest concentration. Mean values for three groups (local herbal, imported regular, imported herbal) have under the permissible limit while two groups (local regular and medicated) have slightly above th
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Mutational Analysis Of Cacna1ggene Implicated In Childhood Absence Epilepsy And Its Comparative Genomics In Mice
Material type: Book ; Literary form:
Publisher: 2017 Dissertation note: Childhood absence epilepsy (CAE) is the subtype of Idiopathic generalized epilepsy (IGE). It accounts for 2-8% of patients with epilepsy. The frequency of CAE is more in girls than boys. The percentage of CAE in youngsters is 10-12%. In addition to CACNA1G, many other genes can be the possible cause of CAE. The pattern of inheritance of CAE is polygenic and complex. SNP might be a gain of function mutation in T- channel genes that results in increase T-type calcium channel activity. Ion channel genes and genes for GABA receptors are affected in epilepsy. By using various techniques of molecular genetics mutations have detected in genes of calcium channels (CACNA1H,CACNA1I, CACNA1A, CACNA1G and CACNB4), in genes of sodium channels like (SCN1B, SCN2A, SCN1A ) and genes for GABA receptor (GABRG2 and GABRD ). Gain of function mutation in CACNA1G gene and increased activity of α1G channels are the possible reason for abnormal SWD in absence epilepsy.
Aim of this study was to assess acknowledged and/or the novel mutations in CACNA1G gene obtained from local childhood epileptic patients.
Blood samples (n=20) were obtained from CAE patients. These samples were collected from children hospital Lahore. Organic method was used to extract DNA from these collected samples. Specific primers were designed for exon 13 and 17 and these exonic regions were amplified using PCR.
After PCR, sequencing of PCR products was performed and then sequencing results were analyzed using chromas lite software.
It has been observed that CACNA1G gene has two mutations in exon 17. It was noticed that protein sequence was altered and the positions of mutations were 38594bp and at 38635bp 38594bp and at 38635 bp. So SNP was detected and there was a gain of function mutation α1G channel activity. In conclusion, these mutations are responsible for absence seizures in CAE patients.
So, it can be concluded that to find out how individuals get affected by these mutations and what factors are involved in causing such mutation, a large scale study should be conducted.In addition, other genes involved in causing epilepsy should also be investigated in local Pakistani Punjab population. As a result of such studies, various diagnostic procedures, strategies for counseling and gene therapies can develop.
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Study On Contamination Of Toxigenic Staphylococcus Aureus In Frozen Desserts And Its Public Health Significance
Material type: Book ; Literary form:
Publisher: 2017 Dissertation note: Frozen desserts are ice cream and other ices which are kept frozen or at low temperature that are said to be frozen dairy desserts. Frozen desserts include ice cream, ice milk and other edible ices. Ice cream is mainly prepared with milk fat which is about 10-16% according to manufacturer standard, without fat milk solids is almost 9-12%, sugar mainly sucrose is about 9-12%, it has water about 55 - 64% and 0.200.50% are stabilizeror emulsifier
In this study 100 samples of commercially prepared frozen desserts were collected and analyzed microbiologically. Nine towns of Lahore were studied, 10 samples were collected from each town from the street venders/local manufacturer, total 90 samples from venders/local manufacturers and 10 samples were collected from the brands outlets. Frozen desserts include dairy frozen dessert, synthetic frozen dessert and some frozen dairy desserts. These samples were culture on Baird Parker agar and the isolated bacteria were further confirmed by PCR tests for detection and identification of the genes encoding Staphylococcal Enterotoxin type A Staphylococcal Enterotoxin type B.
Total 100 samples were processed microbiologically to identify the S. aureuscontamination in frozen desserts. Samples were taken from street vendors and local manufacturers. The results showed that 64% of the dairy frozen desserts were found positive by Staphylococcus spp. 56.25% of the isolates were coagulase positive. The PCR results showed that 17.8% of the S. aureusisolates possess type A gene, while 12.5% had the type B gene, and 9.3% carry both genes.
Laboratory results showed that street vendors which do not have any hygienic conditions as well as temperature maintenance system for their product, they were found contaminated with S. aureus. Major factor of S. aureuscontamination is the personnel handling involvement in the manufacturing frozen desserts. Personnel who were not following hygienic rules to manufacture frozen desserts were found contaminated withS. aureus.S. aureuscan survive at low temperature but cannot grow and produce enterotoxins at low temperature whenever there is a temperature fluctuation S. aureus grows and produce enterotoxins, which could be public health hazard Street vendors don’t have any temperature control system when there is no electricity.Frozen dessert consumers are at risk to get food borne illness or food poisoning.
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