Cloning and sequence analysis of a partial CDS of leptospiral ligA gene in pET-32a - Escherichia coliDH5α system

Research (Published online: 30-04-2018)

23. Cloning and sequence analysis of a partial CDS of leptospiral ligA gene in pET-32a - Escherichia coliDH5α system

Manju Soman, Mangattuparambil Mini, Siju Joseph, Jobin Thomas, Nirmal Chacko, T. G. Sumithra, R. Ambily, Binu K. Mani and Rinsha Balan

Veterinary World, 11(4): 557-561

ABSTRACT

Aim: This study aims at cloning, sequencing, and phylogenetic analysis of a partial CDS of ligA gene in pET-32a - Escherichia coli DH5α system, with the objective of identifying the conserved nature of the ligA gene in the genus Leptospira.

Materials and Methods: A partial CDS (nucleotide 1873 to nucleotide 3363) of the ligA gene was amplified from genomic DNA of Leptospira interrogans serovar Canicola by polymerase chain reaction (PCR). The PCR-amplified DNA was cloned into pET-32a vector and transformed into competent E. coli DH5α bacterial cells. The partial ligA gene insert was sequenced and the nucleotide sequences obtained were aligned with the published ligA gene sequences of other Leptospira serovars, using nucleotide BLAST, NCBI. Phylogenetic analysis of the gene sequence was done by maximum likelihood method using Mega 6.06 software.

Results: The PCR could amplify the 1491 nucleotide sequence spanning from nucleotide 1873 to nucleotide 3363 of the ligA gene and the partial ligA gene could be successfully cloned in E. coli DH5α cells. The nucleotide sequence when analyzed for homology with the reported gene sequences of other Leptospira serovars was found to have 100% homology to the 1910 bp to 3320 bp sequence of ligA gene of L. interrogansstrain Kito serogroup Canicola. The predicted protein consisted of 470 aminoacids. Phylogenetic analysis revealed that the ligA gene was conserved in L. interrogans species.

Conclusion: The partial ligA gene could be successfully cloned and sequenced from E. coli DH5α cells. The sequence showed 100% homology to the published ligA gene sequences. The phylogenetic analysis revealed the conserved nature of the ligA gene. Further studies on the expression and immunogenicity of the partial LigA protein need to be carried out to determine its competence as a subunit vaccine candidate.

Keywords: cloning, Escherichia coli DH5α, Leptospira, ligA, pET-32a, phylogenetic tree.

Prevalence and diversity of gastrointestinal helminths in free-ranging Asian house shrew (Suncus murinus) in Bangladesh

Research (Published online: 30-04-2018)

22. Prevalence and diversity of gastrointestinal helminths in free-ranging Asian house shrew (Suncus murinus) in Bangladesh

Mizanur Rahman, Shariful Islam, Md. Masuduzzaman, Mahabub Alam, Mohammad Nizam Uddin Chawdhury, Jinnat Ferdous, Md. Nurul Islam, Mohammad Mahmudul Hassan, Mohammad Alamgir Hossain and Ariful Islam

Veterinary World, 11(4): 549-556

ABSTRACT

Background and Aim: Asian house shrew (Suncus murinus), a widely distributed small mammal in the South Asian region, can carry helminths of zoonotic importance. The aim of the study was to know the prevalence and diversity of gastrointestinal (GI) helminths in free-ranging Asian house shrew (S. murinus) in Bangladesh.

Materials and Methods: A total of 86 Asian house shrews were captured from forest areas and other habitats of Bangladesh in 2015. Gross examination of the whole GI tract was performed for gross helminth detection, and coproscopy was done for identification of specific eggs or larvae.

Results: The overall prevalence of GI helminth was 77.9% (67/86), with six species including nematodes (3), cestodes (2), and trematodes (1). Of the detected helminths, the dominant parasitic group was from the genus Hymenolepis spp. (59%), followed by Strongyloides spp. (17%), Capillaria spp. (10%), Physaloptera spp. (3%), and Echinostoma spp. (3%).

Conclusion: The finding shows that the presence of potential zoonotic parasites (Hymenolepis spp. and Capillaria spp.) in Asian house shrew is ubiquitous in all types of habitat (forest land, cropland and dwelling) in Bangladesh. Therefore, further investigation is crucial to examine their role in the transmission of human helminthiasis.

Keywords: Asian house shrew, Bangladesh, gastrointestinal helminths, prevalence, Suncus murinus.

Effect of Doublesynch and Estradoublesynch protocols on estrus induction, conception rate, plasma progesterone, protein, and cholesterol profile in anestrus Gir heifers

Research (Published online: 27-04-2018)

21. Effect of Doublesynch and Estradoublesynch protocols on estrus induction, conception rate, plasma progesterone, protein, and cholesterol profile in anestrus Gir heifers

N. J. Chaudhary, D. M. Patel, A. J. Dhami, K. B. Vala, K. K. Hadiya and J. A. Patel

Veterinary World, 11(4): 542-548

ABSTRACT

Aim: This study aimed to evaluate the efficacy of Doublesynch and Estradoublesynch protocols on estrus induction, conception rates, plasma progesterone, protein, and cholesterol profile in anestrus Gir heifers.

Materials and Methods: In this study, 50 pubertal anestrus Gir heifers were selected from the field and farm conditions. The heifers were dewormed (injection ivermectin, 100 mg, s/c) and supplemented with minerals and vitamins (injection organic phosphorus 800 mg and injection Vitamin AD3E and Biotin 10 ml i/m) and multi-mineral bolus at 1 bolus daily for 7 days. The heifers were randomly divided into three groups: Doublesynch (n=20), Estradoublesynch (n=20), and control (n=10). The animals were monitored for estrus response, estrus interval, behavioral signs, and conception rates after induced/first, second, and third cycle post-treatment. Blood samples were obtained on day 0, day 9, day 12, and on day 12 post-artificial insemination (AI) for determination of plasma progesterone, protein, and cholesterol profile.

Results: The estrus response rate between Doublesynch and Estradoublesynch protocols was similar between treated heifers (85% and 95%). The interval from the second prostaglandin F2α (PGF2α) injection to estrus induction did not differ between the groups (63.87±4.19 vs. 58.27±3.83 h). The conception rates following induced estrus (20% vs. 30%), at the second cycle (23.07% vs. 16.66%), at the third cycle (22.22% vs. 30.00%), and the overall conception rate (45% and 55%) within 27.89±5.75 and 26.45±5.48 days were the same across the treatment groups. The mean plasma progesterone concentrations were significantly (p<0.01) higher on day 9 (second PGF2α injection) and day 12 post-AI compared to day 0 (first PGF2α injection) and the day of fixed-timed artificial insemination. The concentrations were also significantly (p<0.05) higher in conceived than non-conceived heifers on day 9 of treatment and day 12 post-AI in both the protocols. The mean plasma cholesterol concentrations were significantly higher during peak follicular and luteal phases compared to the initial anestrus phase in both the protocols. The values were also higher in non-conceived than conceived animals in both the protocols. The plasma protein profile was not influenced by the sampling days or conceived and non-conceived status.

Conclusion: The results showed that both Doublesynch and Estradoublesynch protocols resulted in similar estrus induction and conception rates with modulation of plasma progesterone and cholesterol profile in anestrus Gir heifers.

Keywords: cholesterol, conception rate, estrus synchronization, Gir heifers, progesterone, proteins, pubertal anestrus.

Copy number variation in livestock: A mini review

Review (Published online: 26-04-2018)

20. Copy number variation in livestock: A mini review

V. Bhanuprakash, Supriya Chhotaray, D. R. Pruthviraj, Chandrakanta Rawat, A. Karthikeyan and Manjit Panigrahi

Veterinary World, 11(4): 535-541

ABSTRACT

Copy number variation (CNV) is a phenomenon in which sections of the genome, ranging from one kilo base pair (Kb) to several million base pairs (Mb), are repeated and the number of repeats vary between the individuals in a population. It is an important source of genetic variation in an individual which is now being utilized rather than single nucleotide polymorphisms (SNPs), as it covers the more genomic region. CNVs alter the gene expression and change the phenotype of an individual due to deletion and duplication of genes in the copy number variation regions (CNVRs). Earlier, researchers extensively utilized SNPs as the main source of genetic variation. But now, the focus is on identification of CNVs associated with complex traits. With the recent advances and reduction in the cost of sequencing, arrays are developed for genotyping which cover the maximum number of SNPs at a time that can be used for detection of CNVRs and underlying quantitative trait loci (QTL) for the complex traits to accelerate genetic improvement. CNV studies are also being carried out to understand the evolutionary mechanism in the domestication of livestock and their adaptation to the different environmental conditions. The main aim of the study is to review the available data on CNV and its role in genetic variation among the livestock.

Keywords: copy number variation, copy number variation regions, livestock, single nucleotide polymorphisms, quantitative trait loci.

Molecular detection of Leptospira spp. from canine kidney tissues and its association with renal lesions

Research (Published online: 25-04-2018)

19. Molecular detection of Leptospira spp. from canine kidney tissues and its association with renal lesions

Biswajit R. Dash, Vitthal S. Dhaygude, Prashant D. Gadhave, Kaustubh V. Garud and Dattatarya P. Kadam

Veterinary World, 11(4): 530-534

ABSTRACT

Aim: The study aimed to detect the prevalence of Leptospira spp. in kidney tissues collected during necropsy and to establish its association with renal lesions in dogs of Mumbai region.

Materials and Methods: Kidney tissues from 40 dogs were collected during necropsy after gross examination and then fixed in neutral buffered formalin and Bouin's fluid for histopathology and histochemistry, respectively. Kidney tissues were also collected for the detection of Leptospira spp. by polymerase chain reaction (PCR) in a sterile container and stored at -80°C until further processing.

Results: Of 40 cases studied, 13 (32.5%) cases showed lesions of nephritis of varying histotype and severity. Glomerulonephritis was reported as the most common type of nephritis in 9 (69.23%) cases, and interstitial nephritis was recorded in 4 (30.76%) cases. Chronic and acute interstitial nephritis was observed in two cases each. Renal failure as a cause of death was found in 7 (17.5%) dogs. Of a total of 40 cases, 9 were found positive for pathogenic Leptospira spp. genome by PCR. However, of nine PCR-positive cases, only four cases showed lesions in kidneys as glomerulonephritis and interstitial nephritis in two cases each. The rest five cases positive for Leptospira spp. by PCR did not show any appreciable lesions in the kidneys.

Conclusion: Leptospiral DNA was detected in 9 (22.5%) cases by PCR. Of these nine cases, only four cases showed renal lesions. Other five cases which were positive for Leptospira spp. by PCR did not show any appreciable gross and microscopic lesions in the kidneys which might be carriers for Leptospira spp. Considering variable reports on types of nephritis in Leptospira spp. infection and also the prevalence of non-pathogenic Leptospira spp., it is important to conduct an extensive study on the prevalence of Leptospira spp. and its association with renal lesions involving batteries of tests.

Keywords: histopathology, kidneys, Leptospira spp., nephritis.

Pseudopregnancy in goats: Sonographic prevalence and associated risk factors in Khartoum State, Sudan

Research (Published online: 23-04-2018)

18. Pseudopregnancy in goats: Sonographic prevalence and associated risk factors in Khartoum State, Sudan

Areeg Mohamed Almubarak, Naglaa Abd Elhakeem Abass, Majdi Elnaim Badawi, Mohamed Tagelddin Ibrahim, Abdelhamid Ahmed Elfadil and Rihab Mohamed Abdelghafar

Veterinary World, 11(4): 525-529

ABSTRACT

Aim: This study was conducted to estimate the prevalence of pseudopregnancy in goats and to investigate potential risk factors associated with the condition in Khartoum State.

Materials and Methods: A cross-sectional study was carried out from March 2015 to February 2016. A total of 378 female goats which presented to the Veterinary Teaching Hospital, College of Veterinary Medicine, Sudan University of Science and Technology, for routine ultrasonographic pregnancy diagnosis were examined. Ultrasound scanning was performed using a real-time scanner equipped with dual-frequency (3.5-5 MHz) curvilinear transducer.

Results: The results showed that the prevalence of pseudopregnancy in goats in Khartoum State was 10.6%. Risk factors such as general body condition (χ2=5.974; p=0.05), age (χ2=11.760; p=0.0129), type of estrus (χ2=12.794; p=0.000), and previous reproductive performance (χ2=13.397; p=0.020) showed significant association (p≤0.05) with the occurrence of pseudopregnancy in the univariate analysis. Breed (χ2=12.627; p=0.082), milk yield (χ2=5.951; p=0.114), type of feeding (χ2=1.721; p=0.190), season (χ2=2.661; p=0.264), locality (χ2=7.66; p=0.264), parity number (χ2=0.451; p=0.767), and rearing system (χ2=1.593; p=0.451) were not significantly associated with pseudopregnancy.

Conclusion: The prevalence of pseudopregnancy in goats in Khartoum State was 10.6%. Pseudopregnancy in goats is significantly associated with age, type of estrus, general body condition, and previous reproductive performance. This study showed for the first time that pseudopregnancy is a real reproductive problem in goats in Khartoum State.

Keywords: goat, hydrometra, prevalence, risk factors, Sudan, ultrasound.

Isolation, identification, and serotyping of Avibacterium paragallinarum from quails in Indonesia with typical infectious coryza disease symptoms

Research (Published online: 23-04-2018)

17. Isolation, identification, and serotyping of Avibacterium paragallinarum from quails in Indonesia with typical infectious coryza disease symptoms

Agnesia Endang Tri Hastuti Wahyuni, Charles Rangga Tabbu, Sidna Artanto, Dwi Cahyo Budi Setiawan and Sadung Itha Rajaguguk

Veterinary World, 11(4): 519-524

ABSTRACT

Background and Aim: Infectious coryza (IC) or snot is an infectious upper respiratory disease affecting chickens and birds, including quails, and it is caused by Avibacterium paragallinarum. The symptoms of IC are facial swelling, malodorous nasal discharge, and lacrimation. This study aimed to isolate, identify, and serotype the A. paragallinarum of snot in quails and to determine the sensitivity and resistance to several antibiotics.

Materials and Methods: Nine quails from Yogyakarta, Indonesia with typical snot disease symptoms were used in this study. The nasal swab was obtained and directly streaked onto a chocolate agar plate and blood agar plate (BAP), then incubated in 5% CO2 at 37°C for 24-48 h. Staphylococcus spp. was cross-streaked onto the BAP to show the satellite growth. The observation of the morphology of the suspected colony, Gram staining, and biochemical tests (catalase test, oxidase test, urease test, peptone test, and carbohydrate fermentation such as maltose, mannitol, lactose, and sorbitol) are done to identify the species of bacteria. This research also detects the serovar of A. paragallinarum using hemagglutination inhibition test. The antibiotic sensitivity tests were also performed using several antibiotics against five A. paragallinarum isolates that were cultured on Mueller-Hinton Agar and added with antibiotic discs, then incubated in 5% CO2 at 37°C for 24-48 h.

Results: Five isolates out of nine suspected isolates (55.5%) were A. paragallinarum. The growth of isolates from quails did not depend on the nicotinamide adenine dinucleotide (NAD) (NAD-independent). Sensitivity test was done using the five identified A. paragallinarum isolates, results showed that they were 100% sensitive to amoxicillin (AMC) and ampicillin (AMP); 100% resistant toward amikacin (AK), erythromycin (E), gentamycin (CN), and tetracycline (TE); 80% resistant toward kanamycin (K) and trimethoprim (W); 60% resistant toward chloramphenicol (C); and 20% toward enrofloxacin (ENR). The antibiotics that have an intermediate sensitivity (in between sensitive and resistant) were ENR and K, 80% and 20%, respectively. Three out of five A. paragallinarum isolates were identified as serovar B of A. paragallinarum using HI test.

Conclusion: Five out of nine isolates (55.5%) from quails with typical IC disease symptoms identified as A. paragallinarum and sensitive toward AMC and AMP. Three out of five A. paragallinarum isolates were identified as serovar B.

Keywords: antibiotic sensitivity test, Avibacterium paragallinarum, infectious coryza, nicotinamide adenine dinucleotide-independent.