Morphological and adhesive properties of Klebsiella pneumoniae biofilms

Research (Published online: 28-01-2020)

29. Morphological and adhesive properties of Klebsiella pneumoniae biofilms

Ekaterina Lenchenko, Dmitry Blumenkrants, Nadezhda Sachivkina, Nadezhda Shadrova and Alfia Ibragimova

Veterinary World, 13(1): 197-200

ABSTRACT

Background and Aim: The study of biofilm-forming ability of Gram-negative microflora has great practical importance for assessing the effectiveness of antibiotic therapy and finding new ways to diagnose and inhibit the growth of biofilms. This is because poor penetration of antibacterial drugs into the biofilm can lead to the selection of resistant strains and has a consequence evident by the occurrence of relapse of infection in animals. This study aimed to evaluate morphological and densitometric indicators of biofilm formation as well as adhesive properties of Klebsiella pneumoniae.

Materials and Methods: K. pneumoniae was cultured at 37°C for 2-144 h in vitro. The specimens for optical microscopy were prepared by fixation with a 1:1 alcohol-ether mixture for 10 min and stained with a 0.5% solution of gentian violet for 2 min, and the optical density index was evaluated at a wavelength of 490 nm. Further, the adhesive properties of the microorganisms were determined at a concentration of 1 billion/ml and a suspension of ram erythrocytes at a concentration of 100 million/ml when cultured at 37°C for 24 h. Blood smears were prepared and stained with 0.5% gentian violet.

Results: K. pneumonia cultured at 37°C after 24 h on the meat peptone agar formed large, convex, mucous, and white colonies (d=3.0-6.0 mm). With the growth in the meat and peptone broth, uniform turbidity of the medium was observed. Analyzing the optical density indices (density, D), it was found that K. pneumoniae were good producers of biofilms (D=0.528±0.31). Data for indicators of adhesive properties of K. pneumoniae were as follows: Average adhesion index, 4.56±0.14; adhesion coefficient, 1.07±0.52; and adhesion index, 4.26±0.07. The studied bacteria had high adhesive activity. A direct correlation dependence (R=0.94) of the optical density of biofilms (D≥0.514-0.551) and AAI (4.15±0.28-4.76±0.75) was established.

Conclusion: This study has demonstrated that K. pneumoniae had high adhesive activity, was strong producer of biofilms, and the optical density of the sample exceeded the optical density of the control by more than 4 times.

Keywords: adhesion coefficient, average adhesion index, biofilms, Gram-negative microflora, Klebsiella pneumoniae, optical density.

Protective effect of alpha-lipoic acid and omega-3 fatty acids against cyclophosphamide-induced ovarian toxicity in rats

Research (Published online: 27-01-2020)

28. Protective effect of alpha-lipoic acid and omega-3 fatty acids against cyclophosphamide-induced ovarian toxicity in rats

Dhanya Venugopalan Nair, M. Usha Rani, A. Gopala Reddy, B. Kala Kumar, M. Anudeep Reddy, M. Lakshman and U. Rajkumar

Veterinary World, 13(1): 188-196

ABSTRACT

Background and Aim: Cyclophosphamide therapy is known to be associated with the risk of female infertility as a result of ovarian toxicity. Alpha-lipoic acid (LA) and omega-3 fatty acids are known for their antioxidant and anti-inflammatory activities. The present study investigated the potential protective effect of alpha-LA, omega-3 fatty acids, and its combination against cyclophosphamide-induced ovarian toxicity in rats.

Materials and Methods: Thirty rats were equally divided into Groups I, II, III, IV, and V. Group I was normal control, wherein the rats were fed with normal feed and water ad libitum. Group II served as cyclophosphamide-induced group, wherein the rats were injected with cyclophosphamide at 75 mg/kg through intraperitoneal route once a week to induce ovarian toxicity. Groups III and IV were treated with alpha-LA at the rate of 25 mg/kg and omega-3 fatty acids at the rate of 400 mg/kg, respectively, in parallel to cyclophosphamide induction as in Group II. Group V animals were coadministered with alpha-LA (25 mg/kg) and omega-3 fatty acids (400 mg/kg) along with cyclophosphamide induction as in Group II. The respective treatments were administered daily through oral route for a period of 30 days. Regularity of estrous cycle was evaluated by vaginal cytology. Post-treatment period, the animals were humanely sacrificed, and the blood samples were subjected to the estimation of follicle-stimulating hormone (FSH) and estrogen. The ovarian tissue was weighed and subjected to histopathology, transmission electron microscopy, estimation of decreased glutathione (GSH), and tumor necrosis factor (TNF)-alpha.

Results: Rats treated with cyclophosphamide alone manifested irregularity in estrous cycle, increased FSH, and reduced estrogen levels. The ovaries showed decreased GSH and increased TNF-alpha concentrations. Histopathological and transmission electron microscopic analysis of the ovarian follicles revealed degenerative changes. Administration of alpha-LA and omega-3 fatty acids as well as the combination of both the treatments demonstrated significant normalization of the estrous cycle and antioxidant defense mechanism as well as ameliorated the hormonal profile and histological architecture of the ovarian follicles. However, appreciable synergistic efficacy of the combination therapy (alpha-LA+omega-3 fatty acids) with respect to the monotherapies was not observed in the present study.

Conclusion: The efficacy of alpha-LA and omega-3 fatty acids against cyclophosphamide-induced ovarian toxicity could be attributed to its antioxidant and anti-inflammatory activities that prevented the oxidative damage to the ovaries caused by cyclophosphamide. Hence, our findings suggest that dietary supplementation of alpha-LA and omega-3 fatty acids in women receiving cyclophosphamide therapy could carry potential benefits in preventing cyclophosphamide-induced infertility in childbearing women.

Keywords: alpha-lipoic acid, cyclophosphamide, omega-3 fatty acid, ovarian toxicity, oxidative stress.

Salmonella Enteritidis reduction in layer ceca with a Bacillus probiotic

Research (Published online: 25-01-2020)

27. Salmonella Enteritidis reduction in layer ceca with a Bacillus probiotic

Paul T. Price, Thomas A. Gaydos, Roy D. Berghaus, Virginia Baxter, Charles L. Hofacre and Michael D. Sims

Veterinary World, 13(1): 184-187

ABSTRACT

Background and Aim: Salmonella Enteritidis (SE) is a significant foodborne pathogen that can often be traced to poultry and poultry products. This study aims to evaluate the ability of three commonly used non-antimicrobial feed additives in reducing the amount of SE in the ceca of laying type pullets.

Materials and Methods: On day 0, 60 Hy-Line Brown pullets aged 9 weeks were allocated to individual cages in 15 replicate blocks of four pens. Pullets were administered a mash feed provided ad libitum without supplementation (control) or with dietary supplementation of 454 g/ton yeast cell wall (YCW), or 454 g/ton Bacillus spp. probiotic, or 1133 g/ton yeast culture (YC). On day 3 of the trial, all birds were orally administered 3×107 CFU of a nalidixic acid-resistant SE. On day 10, 7 days after inoculation, all birds were humanely euthanized, and the ceca were aseptically removed for analysis.

Results: There was no significant difference in the prevalence of SE among treatments. The mean quantity of SE detected in the ceca expressed in log10 most probable number/g was 2.52 in the control, 2.49 in the YCW treatment, 1.73 in the probiotic treatment, and 1.66 in the YC treatment. The reduction between control and probiotic and control and YC was significant (p=0.021).

Conclusion: This study demonstrates the ability of the novel probiotic and the YC to reduce the load of SE in layer ceca.

Keywords: layers, probiotic, Salmonella Enteritidis, yeast cell wall, yeast culture.

Toxicity and anthelmintic efficacy of chitosan encapsulated bromelain against gastrointestinal strongyles in Small East African goats in Kenya

Research (Published online: 25-01-2020)

26. Toxicity and anthelmintic efficacy of chitosan encapsulated bromelain against gastrointestinal strongyles in Small East African goats in Kenya

Shukuru Wasso, Naomi Maina and John Kagira

Veterinary World, 13(1): 177-183

ABSTRACT

Background and Aim: The development of resistance to anthelmintic drugs has prompted research into alternative methods of controlling intestinal nematodes in ruminants. This study aimed at evaluating the in vitro and in vivo anthelmintic efficacy and toxicity of chitosan encapsulated bromelain in Small East African goats in Kenya.

Materials and Methods: Adult mortality assay was performed using live Haemonchus contortus worms treated with encapsulated bromelain solution ranging from 0.125 mg/ml to 2 mg/ml. Percentage mortality of worms was calculated after 24 h and the lethal concentration 50% (LC50) determined. For the in vivo study, 18 healthy male indigenous goats were divided into six groups of three goats each. The encapsulated bromelain was orally administered in increasing dosages (3-30 mg kg) once daily, for 14 days. The packed cell volume (PCV), aspartate aminotransferase (AST), alanine aminotransferase (ALT), urea, creatinine, and fecal egg count (FEC) were determined on a weekly basis. At the end of the study, the goats were sacrificed and gross pathology and histopathology of main organs assessed.

Results: Albendazole had the highest (p<0.05) anthelmintic effect on the worms. An LC50 of 0.05 mg/ml, 0.445 mg/ml, and 0.155 mg/ml was observed for albendazole, plain bromelain, and encapsulated bromelain, respectively. The PCV of treated and untreated goats did not show any significant difference (p>0.05), varied from 29.3% to 35.1%, and was within the normal range of the animal. Likewise, no significant differences (p>0.05) were observed between the AST, ALT, urea, and creatinine levels of treated and the control (non-treated) goats. No adverse clinical symptoms, toxicity of the main organs, and mortality in goats were associated with the chitosan encapsulated bromelain after administration of dose up to 30 mg/kg for 14 days. Therefore, the lethal dose 50 of encapsulated bromelain may be considered to be >30 mg/kg. On day 28 post-treatment, the encapsulated bromelain showed a higher in vivo FEC reduction (68.8%) as compared to the plain bromelain (32.4%).

Conclusion: Our results show that bromelain encapsulated in chitosan may be safe and effective in reducing the burden of gastrointestinal tract strongyle nematodes in goats. However, there is a need for further studies to establish the dosage of the encapsulated bromelain to be administered in a single dose for the treatment of goats against gastrointestinal strongyles. In addition, species-specific studies on the efficacy of encapsulated bromelain on strongyles are necessary to evaluate its effectiveness against the entire Strongyloididae family.

Keywords: anthelmintic efficacy, bromelain, chitosan, goats, nanoencapsulation, toxicity.

Multilocus analysis of Gallid herpesvirus 1 in layer chickens in Iraq

Research (Published online: 24-01-2020)

25. Multilocus analysis of Gallid herpesvirus 1 in layer chickens in Iraq

Mohammed Hamzah Abdulkadhim Al-Saadi

Veterinary World, 13(1): 170-176

ABSTRACT

Background and Aim: Infectious laryngotracheitis virus (ILTV) causes a highly pathogenic respiratory disease that affects poultry. It is also known as Gallid herpesvirus 1. ILT prophylaxis measures often include using live attenuated vaccines. The live attenuated vaccine can, however, lead to the formation of new strains of ILTV as a result of vaccine reversion and recombination with field strains. Therefore, this study was performed to explore the multilocus variation of ILTV strains of field and vaccine origin. Samples were tested from two distinctive geographical areas in Iraq as little is known about the ILTV genetic diversity within these areas.

Materials and Methods: The polymerase chain reaction method was utilized to generate sequencing templates of six highly polymorphic genes, including UL54, UL52, gB, ICP18.5, ICP4, and gJ in the layer chicken sample (n=15). The Western blotting technique was also employed to detect and estimate the native molecular weight of gE.

Results: The results revealed an important degree of genetic relatedness between the field and vaccine strains across all genes. In addition, gE was found to be expressed natively at 49 kDa.

Conclusion: The findings of this study may be used to improve the production process of the vaccine for more effective ILT prophylaxis and could further the understanding of epidemiologists and immunologists to better control ILT in the future.

Keywords: Gallid herpesvirus 1, infectious laryngotracheitis virus, infectious laryngotracheitis, polymerase chain reaction.

Effect of immunoglobulin G concentration in dairy cow colostrum and calf blood serum on Cryptosporidium spp. invasion in calves

Research (Published online: 24-01-2020)

24. Effect of immunoglobulin G concentration in dairy cow colostrum and calf blood serum on Cryptosporidium spp. invasion in calves

Alīna Derbakova, Maksims Zolovs, Dace Keidāne and Žanete Šteingolde

Veterinary World, 13(1): 165-169

ABSTRACT

Aim: The research aimed to test the association between the level of immunoglobulin G (IgG) in bovine colostrum and calf blood serum and to evaluate its relation to Cryptosporidium spp. invasion in calves.

Materials and Methods: Fresh colostrum and fecal specimens from cows (n=114) as well as blood and fecal specimens from newborn calves (n=114) were collected in the dairy cattle farm. Investigated calves were separated from their mothers directly after birth and received 2 L of colostrum in two separate feedings within the first 24 h. Blood samples were taken from calves at the age of 2 days. Coprological samples were taken from calves at the age of 1, 10, and 15 days. Both colostrum and fecal samples from cows were taken on the 1st day after calf birth. Rectal fecal samples were collected separately from each calf and cow into plastic bags. The collected calf serum samples and bovine colostrum samples were tested for bovine IgG by competitive enzyme-linked immunosorbent assay kit bovine Ig. To record oocysts of Cryptosporidium spp. in feces, the flotation method was used. Binomial logistic regression was performed to ascertain the effects of IgG in bovine colostrum and calf blood serum on the likelihood of Cryptosporidium spp. infection in calves.

Results: The concentration of IgG in bovine colostrum was higher (70.7±26.6 g/L, mean±standard deviation) than that in calf blood serum (13.2±6.1 g/L); the statistically significant difference was 57.4 g/L (95% confidence interval, 52.4-62.4), t (124.872)=22.536, p<0.001. Mann–Whitney's U-test showed a significant difference between samples collected on days 10 and 15 of the experiment (U=1944, z=2.330, p=0.020). The higher number of oocysts in calf feces was recorded on day 15 (median=6.5) compared to day 10 (median=4). The prevalence of calf infection from days 10 to 15 increased from 26.3 to 45.6% and was at least 3 times higher than in cows. A statistically significant positive correlation was recorded between IgG concentration of cow colostrum and calf blood serum (r (114)=0.414, p=0.001), whereas a correlation between the concentration of IgG and the intensity of Cryptosporidium spp. infection was not recorded (p>0.05). The logistic regression model was not statistically significant (χ2(2)=0.013, p=0.99 (10 days) and χ2(2)=0.100, p=0.95 (15 days)).

Conclusion: Mother passive transfer of immunity to the offspring through colostrum does not influence the susceptibility of calves to Cryptosporidium infestation.

Keywords: calves, Cryptosporidium, dairy cows, immunoglobulins.

The influence of duration on pain stress, oxidative stress, and total antioxidant power status in female dogs undergoing ovariohysterectomy

Research (Published online: 23-01-2020)

23. The influence of duration on pain stress, oxidative stress, and total antioxidant power status in female dogs undergoing ovariohysterectomy

Kanissarinn Sakundech, Chayanon Chompoosan, Pongsatorn Tuchpramuk, Thongchai Boonsorn and Worapol Aengwanich

Veterinary World, 13(1): 160-164

ABSTRACT

Background and Aim: Ovariohysterectomy (OHE) is a common procedure for sterilization of female dogs. However, knowledge of changes in pain stress, oxidative stress, and total antioxidant power status before, during, and after OHE is limited. The objective of this experiment was to study the effect of duration on pain stress, oxidative stress, and total antioxidant power status in female dogs undergoing OHE.

Materials and Methods: Seven female dogs were sterilized using the OHE method. Pain scores, hematological changes, and biochemical markers were investigated during pre-operative, 3 h after starting OHE, and on days 3, 7, 10, and 14 of an experimental period. Data were analyzed using one-way analysis of variance.

Results: At 3 days after OHE, pain score was higher than on days 7-14 of the experimental period; percentage of neutrophil, 3 h after starting OHE, was higher than during pre-operative and on days 3-14 of the experimental period; percentage of lymphocyte on days 10-14 was lower than during pre-operative, 3 h after starting OHE, and on days 3-7 of the experimental period; neutrophil/lymphocyte ratio, 3 h after starting OHE, was higher than during pre-operative and on days 3-14 of the experimental period; plasma malondialdehyde on day 3 was higher than during pre-operative, 3 h after starting OHE, and on days 3-14 of the experimental period; and total antioxidant power on day 14 was higher than during pre-operative, 3 h after starting OHE, and on days 3-10 of the experimental period, respectively.

Conclusion: This experiment indicated that OHE caused pain stress, oxidative stress, and reduction of total antioxidant power in female dogs. Finally, female dogs needed antioxidant for 7 days after OHE.

Keywords: female dog, ovariohysterectomy, oxidative stress, pain stress, total antioxidant power.