Evaluation of a modified method of extraction, purification, and characterization of lipopolysaccharide (O antigen) from Salmonella Typhimurium

Research (Published online: 05-11-2020)

6. Evaluation of a modified method of extraction, purification, and characterization of lipopolysaccharide (O antigen) from Salmonella Typhimurium

Heba M. Hassan, Mai A. Fadel and Mohamed A. Soliman

Veterinary World, 13(11): 2338-2345

ABSTRACT

Background and Aim: Lipopolysaccharide (LPS) is an integral part of the outer cell membrane complex of Gram-negative bacteria. It plays an important role in the induction and stimulation of the immune system. Various LPS purification protocols have been developed. However, analysis of their efficacy is limited by contamination during downstream applications or the public health hazard of LPS. The aim of this study was to evaluate a modified method for extracting LPS as well as assess the purity of the extracted LPS by high-performance liquid chromatography (HPLC) analysis. Further, we evaluated its immunopotentiating function by measuring the relative RNA expression levels of splenic immune-related genes such as interleukin 1β (IL-1β) and interferon-γ (IFN-γ), after intramuscular injection of increasing concentrations of the extracted LPS in specific pathogen-free (SPF) chick.

Materials and Methods: Isolation, identification, and serotyping of Salmonella Typhimurium were performed using chicken flocks. We then performed molecular typing of Salmonella isolates using conventional polymerase chain reaction (PCR). A new protocol for purification of LPS from Salmonella isolate (S. Typhimurium) was conducted. HPLC analysis of the extracted LPS in the current study was compared to existing methods. An in vivo study was performed to evaluate the ability of LPS to induce an immune response by measuring relative IFN-γ and IL-1β gene expression after injecting increasing concentrations of the extracted LPS into SPF chicks.

Results: Isolation and serotyping revealed that Salmonella enterica was of the serovar Typhimurium. Confirmation was conducted by molecular typing through conventional PCR. Fractionation of the LPS extract by HPLC revealed a high degree of purity comparable with standard commercial LPS. These results demonstrate the high purity of extracted LPS based on our modified method using propanol and sodium hydroxide mixture. Intramuscular injection of the extracted LPS in 22 day-old SPF chicks, compared to the negative control, revealed significant upregulation of IFN-γ and slight downregulation of IL-1β.

Conclusion: The new modified method can be used for high purity LPS extraction and demonstrates effective immunopotentiating activity.

Keywords: extraction, high-performance liquid chromatography, interferon-γ and interleukin 1β genes expression, LPS (O antigen), purification, quantitative polymerase chain reaction, Salmonella Typhimurium.

Effects of steroid growth promoter on morphological and biochemical adaptations in liver of broiler

Research (Published online: 05-11-2020)

5. Effects of steroid growth promoter on morphological and biochemical adaptations in liver of broiler

Nasrin Sultana, Marzia Afrose and Kazi Rafiq

Veterinary World, 13(11): 2330-2337

ABSTRACT

Aim: The study was conducted to observe the effects of dexamethasone (DEX) on the gross study and histomorphometry of liver and on the alterations of biochemical parameters of broilers.

Materials and Methods: Ninety day-old chicks were collected and assigned to one of three groups: The control, Group A, and Group B. The control, Group A, and Group B were fed for 28 days with a homemade ration, a commercial broiler type ration, and a homemade ration with DEX (7 mg/kg feed), respectively. Liver samples were collected from the individual birds after sacrificing on days 7, 14, 21, and 28 of the experiment. Morphometric characteristics (length, weight, color, and texture) of the liver were examined. Histomorphological alterations of the liver were assessed with routine hematoxylin and eosin staining. To measure the biochemical parameters, blood samples were collected on days 7, 14, 21, and 28 of the experiment. Liver function test was performed spectrophotometrically by analyzing serum biochemical markers, that is, alanine aminotransferase (ALT), aspartate aminotransferase, and alkaline phosphatase (ALP). Thin-layer chromatography (TLC) was performed for the detection of hepatic steroids.

Results: Hemorrhagic and congested livers were found in broilers of Group B. There were no significant changes found in weight and length of the livers; only numerical decrease in weight and length was observed in birds of Group B. Liver width was increased in Group B on day 21. Histological observation of livers showed accumulation of lipid droplets, congestion of the sinusoids, and central veins in broilers of Group B. Biochemical analyses showed increased levels of ALT in Group B as compared to Group A on day 14 of the experiment. TLC evaluation revealed a positive result in Group B on 28 days of the experiment.

Conclusion: The present study results show that DEX may alter the liver morphology and the concentration of ALT in the circulation of broilers.

Keywords: broiler, dexamethasone, growth promoter, liver.

Seroprevalence of canine hepatitis in stray dogs in Nineveh Province, Iraq

Research (Published online: 04-11-2020)

4. Seroprevalence of canine hepatitis in stray dogs in Nineveh Province, Iraq

Zahraa Mustafa Al-Jumaa, Eva Aisser Ajaj and Mohammad Osamah Dahl

Veterinary World, 13(11): 2326-2329

ABSTRACT

Aim: The current study was conducted to explore evidence of hepatitis B virus (HBV) infection in dogs in Nineveh Province, Iraq.

Materials and Methods: Serum samples of 78 dogs were used to (i) estimate levels of the antibodies against HBV through enzyme-linked immunosorbent assay and (ii) measure the activity of liver function enzymes.

Results: Seropositive dogs for HBV constituted 9% of total tested dogs. The differences in seropositivity among males compared to females and among different ages were not statistically significant. Liver function enzymes analysis revealed a significant increase in the activity of alanine aminotransferase, aspartate aminotransferase, and alkaline phosphatase in seropositive dogs compared to those seronegative.

Conclusion: Hepatitis B is evident in dogs in Nineveh Province, Iraq, with a significant impact on liver function in affected dogs. It is important to confirm this evidence through molecular tests.

Keywords: alanine aminotransferase, alkaline phosphatase, aspartate aminotransferase, dogs, Hepatitis B virus.

Genetic characterization and phylogenetic study of Indonesian cuscuses from Maluku and Papua Island based on 16S rRNA gene

Research (Published online: 04-11-2020)

3. Genetic characterization and phylogenetic study of Indonesian cuscuses from Maluku and Papua Island based on 16S rRNA gene

Rini Widayanti, Richo Apriladi Bagas Pradana, Rony Marsyal Kunda and Suhendra Pakpahan

Veterinary World, 13(11): 2319-2325

ABSTRACT

Background and Aim: Indonesian cuscuses are now becoming scarce because of the reduction of habitat and poaching. Further, molecular characterization of Indonesian cuscuses is still very lacking. This study aimed to determine genetic markers and phylogenetic relationships of Indonesian cuscuses based on 16S rRNA gene sequences.

Materials and Methods: This study used 21 cuscuses caught from two provinces and 16 islands: 13 from Maluku and eight from Papua. Cuscus samples were taken by biopsy following ethics guidelines for animals. The genome isolation was done using gSYNC DNA Mini Kit (Geneaid Biotech Ltd., Taiwan). The 16S rRNA gene was amplified by primers (16SKUSAF and 16SKUSAR), and the polymerase chain reaction product obtained was 1875 base pair (bp). The analysis of genetic characterization and the phylogenetic relationship was performed using MEGA version X software (https://www. megasoftware.net/).

Results: 16S rRNA gene sequencing attained 1598 bp for all samples. Based on the 16S rRNA nucleotide sequences, cuscuses from Papua and Maluku belong to the genus Phalanger and Spilocuscus. Phalanger spp. and Spilocuscus spp. from Papua can be distinguished from Phalanger and Spilocuscus from Maluku, except Spilocuscus from Ternate has a very close relationship with cuscus from Sentani, Papua.

Conclusion: Indonesian cuscuses were derived into two clades based on 16S rRNA gene sequence, one group to genus Phalanger and another group to Spilocuscus.

Keywords: Indonesian cuscuses, Maluku, Papua, Phalanger, phylogeny, Spilocuscus.

Serotypes and antimicrobial resistance profiles of Salmonella enterica recovered from clinical swine samples

Research (Published online: 03-11-2020)

2. Serotypes and antimicrobial resistance profiles of Salmonella enterica recovered from clinical swine samples

Siriporn Kongsoi, Suksun Chumsing, Darunee Satorn and Panisa Noourai

Veterinary World, 13(11): 2312-2318

ABSTRACT

Background and Aim: Salmonella enterica is an important foodborne pathogen and is recognized as a major public health issue. The emergence of multidrug-resistant (MDR) S. enterica represents a major challenge for national public health authorities. We investigated the distribution of serovars and antimicrobial resistance of S. enterica isolates from clinical swine samples stored at the Veterinary Diagnostic Laboratory, Faculty of Veterinary Medicine, Kasetsart University from 2016 to 2017.

Materials and Methods: Clinical samples were collected and subjected to standard microbiological techniques outlined in the Manual of Clinical Microbiology to identify Salmonella serovars. Susceptibility to antimicrobials was tested by the Kirby–Bauer disk diffusion method using a panel of 14 antimicrobials.

Results: A total of 144 Salmonella isolates were identified and the dominant serovar was Salmonella Choleraesuis (66.67%), followed by monophasic Salmonella Typhimurium (18.75%), S. Typhimurium (9.03%), and Rissen (5.56%). The isolates displayed high resistance rates to ampicillin (AMP [100%]), amoxicillin (AX [100%]), tetracycline (TE [100%]), cefotaxime (CTX [89.58%]), ceftriaxone (CRO [87.50%]), chloramphenicol (C [82.64%]), gentamicin (CN [79.17%]), nalidixic acid (NA [72.92%]), and ceftazidime (CAZ [71.53%]). All isolates were MDR, with 29 distinct resistance patterns. The dominant MDR pattern among serovars Choleraesuis and Rissen exhibited resistance to 9 antimicrobials: ( R7-14 AMP-AX-CAZ-CRO-CTX-NA-C-CN-TE). However, all tested isolates were susceptible to AX/ clavulanic acid and fosfomycin.

Conclusion: High resistance levels to the third generation of cephalosporins such as CAZ, CRO, and CTX highlight the need for careful and reasonable usage of antimicrobials in animals and humans, especially for S. Choleraesuis infections.

Keywords: clinical isolates, multidrug resistance, Salmonella Choleraesuis.

Prolactin gene polymorphisms and associations with reproductive traits in Indonesian local ducks

Research (Published online: 03-11-2020)

1. Prolactin gene polymorphisms and associations with reproductive traits in Indonesian local ducks

Dattadewi Purwantini, R. Singgih Sugeng Santosa, Setya Agus Santosa, Agus Susanto, Dewi Puspita Candrasari and Ismoyowati Ismoyowati

Veterinary World, 13(11): 2301-2311

ABSTRACT

Background and Aim: Reproductive traits play an important role in population increases and the egg production (EP) abilities of Indonesian local ducks (ILD). The prolactin (PRL) gene is a single chain polypeptide hormone belonging to a family of growth hormone genes that are mainly synthesized in the anterior pituitary gland in all vertebrates. It has a significant effect on reproductive traits and EP. Single nucleotide polymorphisms (SNPs) present in PRL are a useful molecular marker for EP. This study aimed to identify the PRL polymorphisms based on these SNPs and to uncover the associations with reproductive traits in ILD.

Materials and Methods: A total of 280 ILDs consisting of Tegal and Magelang (F0) ducks and their reciprocal crosses, namely, Gallang (F1) and Maggal (F1), were maintained and specific variables were recorded, that is, age at first egg, body weight at first egg, first egg weight, and EP, for 90 days. Allele and genotype frequencies were used to determine the Hardy- Weinberg (H-W) equilibrium. The association between the SNP genotypes of PRL and reproductive traits was analyzed using one-way analysis of variance, following the GLM procedure of SAS. The genotypic effects on the reproductive traits were determined using regression analysis.

Results: This study successfully amplified a polymerase chain reaction product of 190 bp, which was used to identify the SNP. Results indicated that PRL in ILDs is polymorphic. A SNP was found at position 164 nt (c.164G >A), consisting of three different genotypes, namely, GG, GA, and AA. The genotypes of Tegal and Magelang (F0), and Gallang (F1) populations were not in H-W equilibrium. The Maggal population (F1) was in H-W equilibrium. Significant associations were detected between the genotypes and EP in all ILDs (p<0.01), following a regression line of y=2.337x+64.605, with a determination coefficient of 0.0188 (r=0.14).

Conclusion: PRL can be recommended as a candidate gene for reproductive traits in ILD, especially EP.

Keywords: egg production, Magelang, Prolactin gene, reciprocal mating, single nucleotide polymorphisms, Tegal.

Molecular detection and prevalence of SARS-CoV-2 during the early outbreak in Southern Bangladesh

Research (Published online: 03-11-2020)

10. Molecular detection and prevalence of SARS-CoV-2 during the early outbreak in Southern Bangladesh

Eaftekhar Ahmed Rana, Nahida Sarwer Chowdhury, Md. Sirazul Islam, Jahan Ara, Syeda Shamima Nasrin, Pronesh Dutta, Sabiha Zarin Tasnim Bristi, Tanvir Ahmad Nizami, Prashanta Chakraborty and Amam Zonaed Siddiki

International Journal of One Health, 6(2): 153-159

ABSTRACT

Background and Aim: Coronavirus disease (COVID-19) has been announced as a life-threatening, highly transmissible infectious novel emerging disease worldwide. Rapid detection and epidemiological information are desperately needed to overcome the existing pandemic situation and alleviate national and international crises. Still, to date, there is no significant epidemiological study of COVID-19 available in Bangladesh, especially in the Chattogram division. However, the current study focuses on molecular detection, prevalence, and risk factors associated with COVID-19 in Southern Bangladesh.

Materials and Methods: Standard diagnostic protocols through real-time reverse transcriptase-polymerase chain reaction (RT-qPCR) were performed for molecular confirmation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Different patient demographics were analyzed for exploring the relationship of four factors – region, sex, age, and symptoms with the accumulated number of COVID-19 cases in the Southern Bangladesh during the period of May 13, 2020, to June 12, 2020.

Results: A total of 2954 samples were tested where the cumulative prevalence of circulating SARS-CoV-2 was 29.76% (n=879; 95% CI: 28.11-31.44) in the selected study region. Among the risk factors, the present study revealed that flatland people (35.62%, 95% CI 33.61-37.67, OR=3.13) were more vulnerable to getting infected by SARS-CoV-2 than the people living in hill tracts (13.04%, 95% CI 10.73-15.63). People older than 50 years (34.68%, 95% CI 30.38-39.18) were designated the highest risk than other different age groups. A higher number of COVID-19 cases were confirmed in patients (36.0%, 95% CI 33.77-38.29, OR=1.76) with typical symptoms, but interestingly a significant number of asymptomatic carriers (20.39%, 95% CI 18.13-22.80) also positive for SARS-CoV-2.

Conclusion: To the best of our knowledge, this is the first epidemiological report in the Southern Bangladesh for COVID- 19. The study's findings will contribute to health professionals and the policy-makers to take preventive measures against the next outbreak emergency for Bangladesh.

Keywords: asymptomatic carrier, coronavirus disease 2019, prevalence, risk factors, severe acute respiratory syndrome coronavirus 2.