1.
Molecular Detection And Speciation Of The Canme Piropiasm
by Isma Nazli Bashir | Prof. Dr.Zrafar Iqbal Ch | Dr.Peter J.Irwin | Prof. Dr. Azhar Maqbool | Faculty of Veterinary Sciences.
Material type: ; Format:
print
Publisher: 2008Dissertation note: An epidemiological study of babesiosis in dogs was conducted at Pet center, University of Veterinary and Animal Sciences, Lahore, for one year and information on age, sex and breed was gathered. It was found that from a total number of 6204, dogs up to two years of age were more susceptible than other age groups (2-4, 4-6 and above 6 years).The data regarding genders revealed that males were more prone to the disease than female dogs. As far as the breeds were concerned, crossbred dogs were more susceptible followed by Pointers, Alsatians, German shepherds and Bull terriors.Hot and humid months (June to September) have greater impact on the occurrence of disease. The study regarding identification of ticks revealed that Rhiphicephalus sanguinus is the predominant vector of the disease in Pakistan.
Molecular studies were conducted to characterize and identify the species responsible for canine babesiosis in Pakistan. In this regard, a nested polymerase chain reaction-Restriction fragment length polymorphism (PCR-RFLP) technique was employed on different specimens (Blood, Body tissues and Ticks). For this purpose blood samples were collected from twenty four chronically infected dogs and applied on the Flinders Technologies Associates (FTA) cards for transportation to Australia. Different body tissues (Liver, Spleen, Kidney, Intestine, Bone marrow and Pancreas) were procured after euthanizing the two dogs and DNA was extracted, for further studies. Similarly, the eighty eight ticks were also collected from the infested dogs in the 70% ethanol for transportation to Australia. A nested PCR-RFLP assay was used for the detection and differentiation of Piroplasm species on the basis of the 1 8S ribosomal RNA gene. The assay potentially amplified and identified Babesia gibsoni as the main canine piroplasm. Similar assays on the DNA extracted from body tissues and ticks revealed Babesia gibsoni as the main piroplasm. The PCR was found to have a high detection limit (equivalent to i0 dilution), when using the DNA extracted from blood applied to FTA cards, body tissues and ticks. A new technique was developed for extraction of DNA from FTA cards and tick, in this technique, instead of using the FTA specified punching machine, we used scalpel blades, and so the rest of the chemicals used are'generally and easily available. The same protocol was used for extraction of DNA from ticks, only chemicals used in different quantities with different spinning times. Both of which, resulted in cost reduction, less effort and speedy DNA extraction. The technique reported here has the potential to be standardized for routine DNA extractions from FTA cards and ticks.
Availability: Items available for loan: UVAS Library [Call number: 1061,T] (1).
2.
Serofpidemiology, Zoonotic Potential And Chemotherapy Of Neosporosis In Dogs And Cattle
by Muhammad Mudasser Nazir | Prof. Dr. Azhar Maqbool | Dr. Muhammad Lateef | Prof. Dr.
Material type: ; Format:
print
; Literary form:
drama
Publisher: 2010Dissertation note: The aim of current study was to demonstrate the most important features of Neospora caninum infection in Pakistan. In the present study, I examined the prevalence of N. caninum in 7 districts of the country and to accessed the efficacy of various drugs against the parasite in cell culture.
For the achievement of this purpose, the core objectives were,
To have an overview on the overall seroprevalence of neosporosis throughout the country by means of cELISA in aborting, at risk and clinically healthy cows.
To check the correlation of Iscom ELISA and cELISA, and determination of prevalence of N. caninum by means of Iscom ELISA on milk samples.
To identify the transmission of disease towards human.
To determine the efficacy of various drugs against N. caninum.
In phase 1, overall seroprevalence of N. caninum in dairy cattle (detected by means of cELISA, VMRD, Inc., Pullman, WA, USA) was found to be 43.4% with a significant difference (P < 0.05) of seropositivity among all 18 herds (n = 5 aborting herds, n = 13 non-aborting/clinically healthy herds) selected from 7 districts of Pakistan. The seropositivity of cattle to N. caninum antibodies was significantly higher in aborting animals (52.7%) as compared to non-aborting cows ( 41.5%), indicating a significant difference between aborting and non-aborting cattle. In case of pregnant and non-pregnant animals, similar findings were recorded in our study. A significantly higher rate of seroprevalence was observed in pregnant dams (59.8%) than non-pregnant cattle (35.2%). Overall, higher serological prevalence was evaluated during the summer season (61.1%) in all areas followed by autumn (46.9%), spring (34.9%) and least seropositivity was observed in winter season (26.6%).
The difference in seropositivity was significantly different among all age groups, greater in animals older than 2 years of age. Furthermore, the prevalence was statistically significant (P < 0.05) among cattle of different breeds. Seroprevalence in cases of crossbred animals were higher followed by exotic and indigenous breeds.
Phase 2, describes the seroprevalence of N. caninum in clinically healthy dairy cows. A selection of 760 animals from 13 dairy herds located in Punjab and Sindh Province, Pakistan to demonstrate the presence or absence of the Neospora caninum infection in commercial dairy cattle. The serostatus of the cows towards N. caninum was detected by cELISA (VMRD, Pullman, WA). Out of 760 animals, (43.2%) were seropositive to N. caninum. A significant difference of positivity was recorded among all 13 dairy herds. Age wise prevalence though not statistically significant among all age groups, was greater in animals over 2 years of age and least in heifers. Variation was also observed in samples from cattle of different breeds. A significantly higher prevalence was observed in crossbred animals than in purebred and nondescript cattle. Seasonal prevalence was higher during summer season than rest of the seasons. The seroprevalence of N. caninum in pregnant cows was significantly greater than in non-pregnant animals.
Second experiment of this phase describes the seroprevalence of 240 animals from 5 herds with a high rate of abortion, the percentage of seropositivity observed in these herds was 43.8%, slightly higher than the clinically healthy and non-aborting cattle. No significant difference was observed among all sample locations in this experiment. However, significant difference of positivity was recorded among different breeds of cattle. Age wise prevalence, though not statistically significant (P > 0.05), was greater in animals older than 2 years of age.
The assessment of milk samples from lactating cows were also determined for Neospora caninum antibodies by means of Iscom ELISA (SANOVIR® Sanova Biotech AB, Uppasala, Sweden) and showed a good level of agreement (r² = 0.9959) between the two tests (cELISA and Iscom ELISA). Although, the cELISA (VMRD, Inc., Pullman, USA) expressed a higher seropositivity and sensitivity than Iscom ELISA (Sanova Biotech AB, Uppasala, Sweden). Therefore, both of the ELISA tests (cELISA and Iscom ELISA) for the detection of N. caninum antibodies in dairy cattle can perform better in lactating animals. The Iscom ELISA has some advantages over cELISA as it's easy to collect milk samples than serum samples, moreover Iscom ELISA is cheaper and easy to use but has low sensitivity than cELISA and cannot be used in dry animals. The percentage of positivity detected through Iscom ELISA on individual milk samples were 61.4% and 76.6% by cELISA.
In phase 3, an epidemiological study was conducted to determine seroprevalence of N. caninum in dogs of different breeds and age groups. The serum samples of dogs were analyzed by cELISA (VMRD, Pullman, USA) showed a seropositivity of 23.5%. There was no significant difference of seropositivity among various sample locations, highest prevalence was observed in Muzaffar Garh (31.9%), followed by Gujranwala (27.9%), Lahore (25.1%), Hafizabad (20.2%) and least prevalence was recorded in district Okara (14.6%). A significant difference in prevalence of N. caninum antibodies between male (26.1%) and female (18.8%) dogs were recorded. The difference in seroprevalence was not significant among all age groups. The samples with no age record showed a highest prevalence (29.5%) and least seropositivity was observed in adult dogs of 3-6 years of age (18.7%). During Summer season, highest positivity to N. caninum was (31.0%) recorded while the lowest prevalence (16.0%) was observed in Winter season.
Phase 4, describes the seroprevalence and transmission of N. caninum in humans. A selection of 52 serum samples from humans was analyzed for the presence of N. caninum antibodies. The serostatus of the humans towards N. caninum antibodies (IgG) was determined by using commercially available antigen coated IFAT slides (VMRD, Inc., Pullman, Washington USA) and human conjugate. Overall very low prevalence (1.9%) of N. caninum antibodies was reported in this study. Only one case was found to be positive, these findings indicate that no strong evidence of N. caninum infection in humans.
In phase 5, in vitro drug trials was conducted to access the best efficacy of three commercially available drugs. We found that among three anticoccidial drugs i.e Clindamycin, Diclazuril and Sulfadiazine, Diclazuril has best inhibitory effect against N. caninum tachyzoites in cell culture followed by Clindamycin and sulfadiazine.
Availability: Items available for loan: UVAS Library [Call number: 1575,T] (1).
3.
Epidemiology Zoonotic Potential Haematology Amd Chemotherapy Of Sarcoptic Mange In Camel In Punjab
by Muhammad Irfan Zahid (2011-VA-800 | Prof. Dr. Azhar Maqbool | Prof. Dr. Muhammad Sarwar Khan | Prof. Dr.Shazia Anjum | Prof. Dr. Kamran Ashraf.
Material type: ; Format:
print
; Literary form:
not fiction
Publisher: 2015Dissertation note: A camel is a very hardy ruminant animal, which can survive under harsh climatic
conditions very effectively by utilizing the marginal areas with excellent capabilities and produce
under such conditions (Hjort and Hussein, 1986; Abbas and Tilley, 1990). Camel is an important
animal as it is well adopted in unique manners in the hot, arid and semi-arid environments
(Schwartz, 1992). It can survive without water and food for many days and this unique ability of
camel makes it an ideal for such harsh conditions for which it is also commonly known as “The
Desert Ship”. In spite of the fact that camel is an important member of a group of animals which
produces food for human consumption in the shape of milk and meat, yet it is the most neglected
one in the field of scientific research. It may be due to the fact that camel belongs to such areas
of the world which are arid, semi-arid or rain fed in nature, having harsh climatic conditions,
where poor nutrition and poor management are the major issues (Sohail, 1983).
It is an established fact that diseases originating from parasites lead to the main health
hazard issues in animals. These parasites survive at the expense of the host animals causing lot of
health problems, like skin irritation, anemia leading to weakness and debility. Some of the
parasites have zoonotic importance and may become a source for the transfer of many contagious
diseases like scabies to the human beings (Dominguez et al. 1978). McClain et al. 2009,
observed the scabies as a major health problem globally both for humans and animal population.
Sarcoptes scabiei is an ectoparasite which is a cause of scabies, a skin problem in the human
beings worldwide and the similar species of mites do also produce a similar type of disease in a
large variety of wild and domesticated mammals (Pence and Ueckermann, 2002; Fitzgerald et
al. 2004). Fain, 1978, reported that more than fifteen (15) different species of Sarcoptes scabiei
morphologically and genetically distinct from each other have been identified in different hosts.
Introduction
2
Sarcoptic mange is the second important problematic disease of camel after
Trypanosomiasis (Nayel and Abu-Samra, 1986). Scabies caused by Sarcoptes scabiei var cameli
is a serious & highly contagious skin problem and also economically important disease of the
camels (Pegram and Higgins, 1992). Camels, which are reared with deficient nutrition, poor
management and under unhygienic conditions are mostly affected by this disease (Kumar et al.
1992).
A large group of people and communities living in arid diverse ecozones in the entire
world, particularly in harsh climates earns their livelihoods by depending on camels. This
dependence may spread to the utilization of camel milk, meat, wool and leather besides its use in
transportation, riding and sports (Wilson, 1984; Snow et al. 1992). In Pakistan camels are also
raised by the people for meat, milk, riding, transportation and sports purposes in the deserts, semi
desert & rain-fed / warm areas of the entire country being a hardy animal as it can tolerate easily
the rugged climate as well as extremes of temperatures of such areas.
The natural harsh and adverse climatic conditions, particularly during long dry seasons
lead to a paucity of feeding regimes resultantly the camels raised in such areas are subjected to
stress conditions which lower their resistance and make them easily vulnerable to diseases
(Abbas et al. 1993; Agab, 1993). Abbas & Tilley, 1990; Saint-Martin et al. 1992; Abbas and
Agab, 2002; Pathak and Chhabra, 2010; while reviewing the parasites & parasitic diseases of
camel population in India were of the opinion that Sarcoptic mange is a serious, debilitating,
dreaded and widely prevalent disease of camels in India.
Besides other infectious diseases of bacterial and viral origin, camels are exposed to a
wide range of internal & external parasitic infestations. Amongst other so many external
parasites to which camels are exposed, the Sarcoptic mange is recognized to be one of the most
Introduction
3
serious and damaging disease. This disease is caused by a mite known as Sarcoptes scabiei var
cameli which belongs to genus Camelus of SARCOPTIDAE family in Veterinary Entomology.
It is an extremely pruritic, contagious and debilitating skin disease which is very
frequently and sudden in onset. It is also ranked as one of the most serious and important disease
of the camels. Sarcoptic mange infestation is very common in the areas of thin skin, the head,
neck, flanks, medial aspect of thighs or inguinal region, mammary glands and prepuce. The head
is usually affected very rapidly as the animal uses its teeth for scratching the affected areas.
Besides linking the occurrence of the disease with poor camel management, malnutrition and
contact with infected objects, the stray & infected camels also often become a focus of infecting
the healthy animals when mingling with them particularly at watering places for drinking
purpose (Richard, 1987; Abdel-Rehman et al. 2001).
Sarcoptes is a burrowing mite as it penetrates deeply through the skin surface of the
infected camel. This burrowing of mites in the skin helps these parasites lead to intense pruritus
and exudative dermatitis. In pruritus, mites penetrate deep into muscular areas, damaging the
flesh and lowering the quality of meat. The early inflammatory reaction of the host body towards
the mites becomes evident in the shape of small popular elevations, invasion and injuries leading
to formation of hairless areas, scaly crust formation or scabs on the affected parts and the skin
become dark and thickened. Skin of mangy camel show hemorrhages, and subcutaneous odema
after the development of fissures in the underlying epidermis (Kumar et al. 1992; Amer et al.
2006).
The fertilized female mites create winding burrows or tunnels in the upper layers of the
epidermis of the skin of the host animal and feeding on the serous exudate, a liquid oozing from
the damaged tissues. The female mites lay about 40-50 fertilized eggs in these tunnels which
Introduction
4
hatch in 3-5 days into a six legged larvae. These larvae immediately crawl to the surface and
burrow themselves in the superficial layers of the skin and create small molting pockets. In these
molting pockets, the larvae molt to next stages of nymph and adult. The adult male then emerges
and seeks a female either in the molting pocket or on the surface of skin. After fertilization the
female produces new tunnels, either de novo or, by extension, of the molting pockets, lays eggs
in these tunnels and a new life cycle starts. The entire life cycle of Sarcoptic mange is completed
in 17-21 days.
New hosts can be infected through direct transmission by contact between the animals,
presumably from larvae, nymph or adult mites, which are commonly present on the skin surface
of the infected animal. Indirect transmission of infestation can also take place through the objects
or fomites having mange infection, which come into contact with the affected camel, such as
harnesses, blankets, baggage tack, tents and tree trunks (Richards, 1987). The pruritus increases
as the mites penetrate deeper in the skin (Al-Rawashdeh et al. 2000, Driot et al. 2011, Bekele et
al. 2012). Based on the rate of infection camels can be seriously disturbed by the Sarcoptic
infestation as they may stop grazing which can lead to a rapid fall in milk production, and
deterioration of health condition. With the increase in the irritation due to scabies, the camel
rubs, bites and scratches the affected areas in an attempt to reduce the itchiness. Due to rubbing,
biting or scratching, the mites move to the periphery affecting the healthy tissues and resultantly
affected area spreads. As the disease prolongs, the skin becomes excoriated, leading to hair loss
and the development of scabs. These scabs in turn may be rubbed away and a red surface
developed. The animal becomes restless due to severe Sarcoptic mange infestation and
involvement of most of the body surface. If the diseased animal is not treated in time, the animal
loses its health condition, become emaciated and within two, three weeks the acute stage of
Introduction
5
disease may give way to more chronic state (Gorakh et al. 2000, Abubakar et al. 2002, Driot et
al. 2011). Sarcoptic mites rarely survive long off the host under natural conditions.
A continuous direct contact of animal keepers with their camels can also lead to
transmission of diseased condition in human beings which is termed as pseudo scabies.
Transmission of infection from camel to man usually takes place during milking, handling or
riding. The main symptoms of pseudo scabies can therefore be seen in the inter digital spaces of
the hands, on the wrists, forearms, the elbows, the axillary folds and inner side of the thighs.
Once a herd is infected with Sarcoptic mange, continuous reinfection of the disease occurs
(Schillinger 1987, Singh & Veer 2005, Premalatha et al. 2010).
Sarcoptic mange is usually considered to be a seasonal disease and is often reported
severe during the winter months as in cold weather the disease had an acute course. However,
there is some evidence that in some countries hot weather predisposes to acute outbreaks of
camel mange and in the cooler, winter season the rate of mange infestations are at the lowest. In
the summer the activity of the mite seems to decline or disease becomes chronic. Dietary intake
is an important factor in mange infestation. Nomadic camels on a low nutrition plan, probably
carrying heavy worm burdens in hot desert conditions are likely, therefore, to be highly prone to
Sarcoptes at this time (Dinka et al, 2010). During such periods of great activity, the mites are
readily transmissible from one animal to other animals (Richards, 1987, Banaja & Ghandour,
1994, Tefera & Gebreah, 2001).
Mange can easily be diagnosed clinically from the occurrence of pruritus, depilation,
alopecia, thickened skin, folds around the joints and encrusted plaques being the main
characteristics of this parasitosis. In order to control this zoonotic disease, it is essential to treat
Introduction
6
both camel and man along with effective checks over other predisposing factors of the disease
such as hygiene and nutritional requirements of the animals.
The skin diseases like the scabies both in human beings and animals are being treated
with a variety of allopathic drugs now a day, but the role of herbal plants in use since centuries in
different shapes cannot be ignored at all, especially in the rural lifestyle. Further with the
continuous use of different acaricidal drugs, the issue of resistance development has come across
as a challenge for the researchers to find some alternatives for the purpose. Accordingly the
research work on the use of traditional herbal medicines is gaining attention day by day.
Although there are many reports and studies regarding the prevalence of Sarcoptic mange
in camel from different parts of the world, only few preliminary reports are available for Pakistan
and none of them provide detailed epidemiology of Sarcoptic mange and its effect on host
health. Therefore, keeping in view the importance of the mange problem in camel population of
the country, the present project was designed to determine the prevalence of Sarcoptic mange
infestation, factors in its occurrence its zoonotic importance, effect on blood physiology and
different treatment options in the camel population of Punjab, province in Pakistan. Availability: Items available for loan: UVAS Library [Call number: 2190,T] (1).
4.
Isolation and Identification of Parasites From Available Raw Vegetables
by Sawera Chaudhry (2008-VA-189) | Prof. Dr. Azhar Maqbool | Dr.Sarwat Naz | Dr. Muhammad Latif | Dr. Muhammad Avais.
Material type: ; Literary form:
not fiction
Publisher: 2015Dissertation note: Blank CD. Availability: Items available for loan: UVAS Library [Call number: 2232-T] (1).
5.
Taxonomy And Control Of Flea Infestation In Cats At Lahore
by Umair Tariq (2008-VA-233) | Dr. Nisar Ahmad | Prof. Dr. Azhar Maqbool | Dr. Syed Saleem Ahmad.
Material type: ; Literary form:
not fiction
Publisher: 2015Dissertation note: INTRODUCTION
Fleas play an important role in causing clinical skin disorders and diseases transmission in man
and pets animals (Rust & Dryden, 1997). Fleas are one of the most important ectoparasites with
more than 2,000 species worldwide affecting mammals, birds, and reptiles (Hsu, 2003). In some
locations, fleas represent over 50% of all the dermatological cases presented to small animal
clinics. Most are limited to hosts with nests as this can provide conditions for the completion of
their life cycle (Linardi & de Avelar, 2014). While fleas on pets are generally considered a
nuisance that may cause some dermatologic problems, they are also responsible for the
transmission of several important diseases in humans and animals (Dryden & Rust, 1994). They
have been involved in transmission of cat scratch disease (Bartonella henselae) (Chomel et al.,
2006; Comer et al., 2001), Rickettsia typhi (Murine thyphus), Rickettsia felis (Finkelstein et al.,
2002; Rolain et al., 2005), and also serve as the intermediate host for the tapeworm Dipylidium
caninum (Rust & Dryden, 1997) and several trypanosomatids (Coutinho & Linardi, 2007).
The term ‘‘cat flea,’’ which is the approved common name for Ctenocephalides felis felis (C. f
felis), can occasionally cause confusion. When it appears in print, it refers to the specific flea
genus and species and not to fleas recovered from cats. There are four recognized subspecies of
C. felis throughout the world: Ctenocephalides felis damarensis and C. felis strongylus occur
primarily in East Africa, C felis orientis occurs in India and Australia, and the widespread C. f
felis occurs in all continents except Antarctica and is the only subspecies that occurs in North
America (Rust & Dryden, 1997). The cat flea, C. felis, is a clinically important parasite of
domestic pets, being responsible for the production of allergic dermatitis, serving as the vector of
Introduction
2
various bacterial pathogens, and being the intermediate host for filarid and cestode parasites.
Flea allergy dermatitis is the most common dermatologic disease of dogs and a major cause of
feline miliary dermatitis (Dryden & Rust, 1994; Rust & Dryden, 1997).
Clinical features vary from asymptomatic to severe hypersensitivity reactions with restlessness,
alopecia from scratching and biting resulting in a pruritic papular dermatitis. Vacuuming of
carpets, furniture cushions, rugs, or other substrata, with a vacuum machine containing a ‘‘beater
bar,’’ will remove many of the flea eggs and larvae. In addition, cocooned pupae at the upper
levels of the carpet can also be affected. The vibration also stimulates adult fleas to emerge from
their cocoons so that they can be collected in the vacuum machine. Therefore frequent
vacuuming, during a flea infestation, can reduce the overall flea burden in the home. It should be
ensured that vacuum bags are disposed of properly, to prevent recolonization of the home with
flea stages previously removed by vacuuming. Because outdoor development of immature flea
life stages is limited to shaded areas, altering outdoor environments to eliminate such habitats
can effectively reduce flea populations. Because urban wildlife, such as opossums, raccoons, and
foxes, are good hosts for cat fleas, pet owners should avoid encouraging visitations by wildlife,
which will affect flea and tick control (see later discussion). Treatment of indoor and outdoor
environments with insecticides requires knowledge of what to use and where to use it. For this
reason, it is suggested that pet owners consult with a licensed pest control specialist for such
applications (Angelbeck-Schulze et al., 2014; Perrins & Hendricks, 2007).
In line with increasing urbanization over the last few decades, flea species that infest pets have
become household pests. Thus, and for reasons of animal and human welfare, the control of fleas
is of great importance worldwide. Despite the increase in the number of products available and
Introduction
3
their use, flea infestation of cats and dogs is still widespread in Europe and on other continents,
whereas resistance of these insects against many chemicals has been detected (El-Gazzar et al.,
1986). Cat fleas are the most important ectoparasite of cats and dogs worldwide. During the past
ten years, topical and oral applications of insecticides such as fipronil, imidacloprid, lufenuron
and, most recently, selamectin have revolutionized cat-flea control. Recent studies show that
these therapies eliminate the need to treat indoor and outdoor environments, and their use
markedly reduces the severity and prevalence of flea allergic dermatitis. Surveys have yet to
reveal the development of insecticide resistance to these chemical compounds. Extending the
longevity of these effective host-targeted therapies should be a major goal of the veterinary
community (Rust, 2005). Availability: Items available for loan: UVAS Library [Call number: 2253-T] (1).
6.
Evaluation Of Risk Factors And Molecular Diagnosis Of Dermatophytosis In Dogs
by Muhammad Haseeb Saeed (2008-VA-241) | Prof. Dr. Aneela Zameer Durrani | Dr. Hassan Saleem | Prof. Dr. Azhar Maqbool.
Material type: ; Literary form:
not fiction
Publisher: 2016Dissertation note: Dogs are most kept and beloved pets in Pakistani society. Dermatophytosis is among the common disease of the pets. Many predisposing factors are involved in development of clinical cases of dermatophytosis including climatic conditions, housing condition of dogs and physical attributes such as coat hair size. Dermatophytosis is not only of concern as being infection of pets but also of its zoonotic importance hence it is very crucial to diagnose dermatophytic infection well in time. Dermatophytosis is caused by Dermatophytes,Microsporum, Trichophyton and Epidermophyton, the fungal species. It is difficult to diagnose the Dermatophytosis from other skin infections by routine tests in most of the cases especially subclinical. Polymerase Chain Reaction (PCR) is advanced and the most reliable technique to detect genome of Dermatophytes even in minute quantities specifically and can efficiently detect the presence of any Dermatophyte specie on the skin of dog. The current study was planned to develop and validate a diagnostic assay which could be able to detect and distinguish tree important dermatophytes species including Microsporum, Trichophyton and Epidermophytonby a uniplex PCR reaction. Analysis of involvement of certain predisposing factors in dermatophytosis was second goal to be worked on in this study. Samples of suspected pet dogs (n=50) were collected by scraping the skin at affected areas over skin. DNA was extracted from the skin scraping samples by organic Phenol Chloroform Isoamyle Alcohol method. Primers, specific to the 18-S ribosomal RNA region of genomes of the Dermatophytes, were designed after alignment of available sequences of Microsporum,Trichophyton and Epidermophyton at NCBI. Annealing temperature and recipe of PCR reaction was optimized by gradient PCR in BIO-Rad thermal cycler. Amplification reaction of all samples collected was carried out as per optimized reaction conditions, afterwards. Amplified products obtained were subjected to genotyping by agarose gel electrophoresis for size based separation of the amplified products. The specific amplified bands of desired genomic region of dermatophytes were seen in UV light transilluminator. The data of results of predisposing factors involved in dermatophytosis wasanalysedby using Pearson’s chi squared test with the help of Statistical Package for the Social Science (SPSS) Program.
Genome specific product sizes of Microsporum and Trichophyton i.e. 366 bp and 351 bp in respective positive samples were observed. Out of 50 suspected samples 46 samples were positive for dermatophytosis out of which 38 samples (82.6%) were positive for Microsporum, 6 samples (13%) for Trichophyton and 2 samples (4.4%) were positive for both Microsporumand Trichophyton.
This study will help to validate a diagnostic technique for Dermatophytosis with greater efficacy and reliability. Moreover, this investigation may become basis for the future research activities in this field in Pakistan.
Availability: Items available for loan: UVAS Library [Call number: 2528-T] (1).
