Despite the availability of Newcastle disease (ND) vaccines for more than six decades, disease outbreaks continue to occur with huge economic consequences to the global poultry industry. The aim of this study is to develop a safe and effective inactivated vaccine based on a recently isolated Newcastle disease virus (NDV) strain IBS025/13 and evaluate its protective efficacy in chicken following challenge with a highly virulent genotype VII isolate. Firstly, high titre of IBS025/13 was exposed to various concentrations of binary ethylenimine (BEI) to determine the optimal conditions for complete inactivation of the virus. The inactivated virus was then prepared in form of a stable water-in-oil emulsion of black seed oil (BSO) or Freund's incomplete adjuvant (FIA) and used as vaccines in specific pathogen-free chicken. Efficacy of various vaccine preparations was also evaluated based on the ability of the vaccine to protect against clinical disease, mortality and virus shedding following challenge with highly virulent genotype\VII NDV isolate. The results indicate that exposure of NDV IBS025/13 to 10 mM of BEI for 21 h at 37 °C could completely inactivate the virus without tempering with the structural integrity of the viral hemagglutin-neuraminidase protein. More so, the inactivated vaccines adjuvanted with either BSO- or FIA-induced high hemagglutination inhibition antibody titre that protected the vaccinated birds against clinical disease and in some cases virus shedding, especially when used together with live attenuated vaccines. Thus, genotype VII-based NDV-inactivated vaccines formulated in BSO could substantially improve poultry disease control particularly when combined with live attenuated vaccines.
Village chicken production, a traditional, small-scale, and extensive backyard poultry industry, has been profitable for local farmers in Myanmar. However, there is scanty information available concerning the infection of these chickens with avian pathogens, including haemoprotozoan parasites. In the present study, we provide the first report of microscopic detection and molecular identification of Leucocytozoon and Plasmodium parasites from seven different areas of Myanmar. Leucocytozoon gametocytes were detected in 17.6% (81/461) of the blood smears from village chickens. The nested polymerase chain reaction (PCR) for targeting Leucocytozoon mitochondrial cytochrome b (cyt b) genes had a 17.6% positive rate. Although the positive rate of nested PCR targeting Plasmodium/Haemoproteus cyt b was 34.3%, the PCR protocol was observed to possibly amplify DNA of a certain species of Leucocytozoon. There were no obvious clinical signs in the infected birds. Statistical analysis of the microscopic detection and PCR detection rates using the age and sex of birds as internal factors revealed that the statistical significances differed according to the study area. The sequencing of 32 PCR products obtained from each study area revealed infection by Leucocytozoon caulleryi in three birds, Leucocytozoon sabrazesi in two birds, Leucocytozoon schoutedeni in two birds, Leucocytozoon sp. in eighteen birds, and Plasmodium juxtanucleare in seven birds; however, Haemoproteus infection was not detected. While L. sabrazesi was detected in chickens from the central region of Myanmar, the other haemosporidians were detected in those from different areas. In the haplotype analysis, we detected 17 haemosporidian cyt b haplotypes, including two for L. caulleryi, one for L. sabrazesi, two for L. schoutedeni, nine for Leucocytozoon sp., and three for P. juxtanucleare. Phylogenetic analysis of the cyt b haplotypes revealed a considerably close genetic relationship among chicken haemosporidians detected in Myanmar, Thailand, and Malaysia. These results indicate that well-recognized widespread species of chicken Leucocytozoon and Plasmodium are distributed nationwide in Myanmar, providing new insights into the ecosystem and control strategies of haemosporidian parasites in domesticated chickens in Myanmar.
Polyacrylonitrile nanofiber membrane functionalized with tris(hydroxymethyl)aminomethane (P-Tris) was used in affinity membrane chromatography for lysozyme adsorption. The effects of pH and protein concentration on lysozyme adsorption were investigated. Based on Langmuir model, the adsorption capacity of P-Tris nanofiber membrane was estimated to be 345.83 mg/g. For the operation of dynamic membrane chromatography with three-layer P-Tris nanofiber membranes, the optimal operating conditions were at pH 9, 1.0 mL/min of feed flow rate, and 2 mg/mL of feed concentration. Chicken egg white (CEW) was applied as the crude feedstock of lysozyme in the optimized dynamic membrane chromatography. The percent recovery and purification factor of lysozyme obtained from the chromatography were 93.28% and 103.98 folds, respectively. Our findings demonstrated the effectiveness of P-Tris affinity nanofiber membrane for the recovery of lysozyme from complex CEW solution.
The gastrointestinal tract of chickens harbors a highly diverse microbiota contributing not only to nutrition, but also to the physiological development of the gastrointestinal tract. Microbiota composition depends on many factors such as the portion of the intestine as well as the diet, age, genotype, or geographical origin of birds. The aim of the present study was to demonstrate the influence of the geographical location over the cecal microbiota from broilers. We used metabarcoding sequencing datasets of the 16S rRNA gene publicly available to compare the composition of the Argentine microbiota against the microbiota of broilers from another seven countries (Germany, Australia, Croatia, Slovenia, United States of America, Hungary, and Malaysia). Geographical location played a dominant role in shaping chicken gut microbiota (Adonis R2 = 0.6325, P = 0.001; Mantel statistic r = 0.1524, P = 4e-04) over any other evaluated factor. The geographical origin particularly affected the relative abundance of the families Bacteroidaceae, Lactobacillaceae, Lachnospiraceae, Ruminococcaceae, and Clostridiaceae. Because of the evident divergence of microbiota among countries we coined the term "local microbiota" as convergent feature that conflates non-genetic factors, in the perspective of human-environmental geography. Local microbiota should be taken into consideration as a native overall threshold value for further appraisals when testing the production performance and performing correlation analysis of gut microbiota modulation against different kind of diet and/or management approaches. In this regard, we described the Argentine poultry cecal microbiota by means of samples both from experimental trials and commercial farms. Likewise, we were able to identify a core microbiota composed of 65 operational taxonomic units assigned to seven phyla and 38 families, with the four most abundant taxa belonging to Bacteroides genus, Rikenellaceae family, Clostridiales order, and Ruminococcaceae family.
Stress and fear responses were evaluated in broiler chicks that were pretreated for 24 h with 0 ppm (control) or 1,200 ppm of L-ascorbic acid (AA) in their drinking water. The birds were subsequently subjected to either upright handling (UH) or inverted (IH) handling for about 45 s. Heterophil (H) counts, lymphocyte (L) counts, and H/L ratios (H/L) ratios were determined immediately (T0) and at 20 h (T20) following the handling treatment. The H/L ratios were similar for both groups at T0, whereas 20 h after the handling treatment, AA-supplemented birds had lower ratios than controls, resulting in a significant water treatment x time of blood sampling interaction. Inverted handling had negligible effect on H/L ratios but augmented tonic immobility (TI) durations as compared with UH. Irrespective of handling procedure, supplemental AA reduced underlying fearfulness, as measured by TI reaction. Neither water treatment nor handling method had significant effect on number of attempts to induce TI.
Since 1954, avian mycoplasmosis has been considered a significant problem in chicken flocks in Japan and in other Asian countries. In Japan, Mycoplasma gallisepticum (MG) and M. synoviae (MS) infections were confirmed aetiologically in chicken flocks affected with respiratory disease or synovitis in 1962 and 1973, respectively. In other Asian countries, including Indonesia, the People's Republic of China, Korea, Malaysia, the Philippines, Taipei China and Thailand, the occurrence of mycoplasmosis in chicken flocks has been recognised serologically or aetiologically. Adverse atmospheric and environmental conditions, in addition to mixed infections of bacterial or viral origin, play an important role in the spread of MG and MS within chicken flocks or in the induction of clinical respiratory mycoplasmosis. Serological tests are important in determining and monitoring the mycoplasmal infection status of chicken flocks. The establishment of mycoplasma-free breeding stocks is recognised as essential for the control of avian mycoplasmosis. To eliminate the transmission of MG to the egg, treatment of infected breeder flocks or their progeny with anti-mycoplasmal antibiotics was effective in considerably reducing the infection rate but not in entirely eliminating MG infection. The preincubation heat treatment of chicken hatching eggs has proved an effective procedure for establishing MG- and MS-free breeding stocks in Japan. Vaccination against MG infection has been practised successfully in Japan and other countries.
Single strains of Lactobacillus acidophilus and Lact. fermentum, isolated from chicken intestine, were used to study in vitro interactions with Salmonella enteritidis, Salm. pullorum or Salm. typhimurium in an ileal epithelial cell (IEC) radioactive assay. Exclusion, competition and displacement phenomena were investigated by respectively incubating (a) lactobacilli and IEC together, prior to addition of salmonellae, (b) lactobacilli, IEC and salmonellae together, and (c) salmonellae and IEC, followed by the lactobacilli. Lactobacilli were selected for study because of their strong ability to adhere to IEC and poor aggregation with salmonellae. The results demonstrated that Lact. acidophilus significantly reduced (P < 0.05) the attachment of Salm. pullorum to IEC in the tests for exclusion and competition, but not in the displacement tests. Lactobacillus fermentum was found to have some ability to reduce the attachment of Salm. typhimurium to IEC under the conditions of exclusion (P < 0.08), competition (P < 0.09), but not displacement. However, both Lact. acidophilus and Lact. fermentum were unable to reduce the adherence of Salm. enteritidis to IEC under any of the conditions.
Two Lactobacillus isolates, Lact. acidophilus I 26 and Lact. fermentum I 25, were selected, based on their poor aggregation with Escherichia coli and strong ability to adhere to ileal epithelial cells (IEC), to study in vitro interactions with E. coli O1:K1, O2:K1 and O78:K80 in an IEC radioactive-assay under the conditions of exclusion (lactobacilli and IEC, followed by the addition of E. coli), competition (lactobacilli, IEC and E. coli together) and displacement (E. coli and IEC, followed by the addition of lactobacilli). The results indicated that Lact. acidophilus I 26 and Lact. fermentum I 25 could not significantly reduce the attachment of E. coli O1:K1, O2:K1 and O78:K80 to IEC under the three conditions tested in vitro, except that the attachment of E. coli O1:K1 was slightly reduced by Lact. fermentum I 25 in the test for competition.
Twelve Lactobacillus strains isolated from chicken intestine were used to investigate acid and bile tolerance in vitro. Ten out of the 12 strains were slightly affected by 0.3% bile salts, showing a delay of growth (d) of 0.6-37.2 min compared with growth in control cultures. Two strains were not affected by the bile salts. Of the 12 strains, seven could be arbitrarily classified as resistant (d < 15 min) and five as tolerant (15 min < d < or = 40 min). Lactobacillus strains from the caecum showed better tolerance to acid than those from the ileum. Generally, the survival of the ileal strains was very low at pH 1.0 and 2.0, and moderate at pH 3.0. In contrast, caecal Lactobacillus strains could survive at pH 1.0 for up to 2 h of incubation; growth was moderate at pH 2.0 and good at pH 3.0 and 4.0.
A two-step SYBR Green I real time polymerase chain reaction (PCR, real time PCR) for the detection of Newcastle disease virus (NDV) was developed. A melting curve analysis was performed to distinguish specific from non-specific products and primer dimers. Regardless of different virus pathotypes the melting temperature (Tm) ranged from 86 degrees C to 87 degrees C. The sensitivity of the real time PCR was compared with the reverse transcription (RT)-nested PCR enzyme-linked immunosorbent assay (ELISA, RT-nested PCR ELISA). Whereas the detection limit of the real time PCR was 10 pg DNA, the RT-nested PCR ELISA and conventional PCR could only detect up to 1 ng and 10 ng DNA, respectively. Thus the real time PCR offers a sensitive, rapid and convenient method for screening large number of NDV specimens.
An experimental oil emulsion Newcastle disease vaccine was evaluated for its efficacy in broiler chickens. A group of chickens vaccinated at one day old with a live lentogenic Newcastle disease vaccine and subsequently revaccinated at three and eight weeks old with the experimental oil emulsion vaccine showed satisfactory haemagglutination inhibition antibody response which persisted for 18 weeks. Between 90 and 100 per cent of the vaccinated chickens were protected when challenged with the velogenic viscerotropic Newcastle disease virus. Although the vaccinated chickens were protected against clinical disease, virus could be isolated from a number of birds. By day 10 to 12 after challenge all the chickens were free from Newcastle disease infection.
Meat chickens housed on a commercial broiler farm in Australia were vaccinated once at 10 to 11 days-of-age by aerosol with live V4 Newcastle disease virus (NDV) vaccine. Groups of vaccinated and unvaccinated birds were flown to Malaysia, where they were challenged with a virulent strain of NDV. Survival rates in vaccinated chickens challenged 7, 14, 21 or 31 d after vaccination were 0.47, 0.77, 0.97 and 0.92, respectively. All unvaccinated chickens died due to Newcastle disease (ND) following challenge. Chickens in Australia and Malaysia were bled and the serums tested for haemagglutination-inhibiting (HI) antibody to NDV. Many vaccinated birds with no detectable antibody, and all birds with a log2 titre of 2 or greater, survived challenge. The results showed that this V4 vaccine induced protective immunity in a significant proportion of chickens within 7 d of mass aerosol vaccination. This early immunity occurred in the absence of detectable circulating HI antibody. Non-HI antibody mediated immunity continued to provide protection up to 31 d after vaccination. Almost all vaccinated birds were protected within 3 w of vaccination. It is concluded that the V4 vaccine is efficacious and could be useful during an outbreak of virulent ND in Australia.
The food pellet vaccine has been shown to be effective in trials conducted under laboratory and simulated field conditions. The village chickens vaccinated with the food pellet vaccine during the field trial were protected against virulent Newcastle disease virus. The efficacy of the food pellet vaccine in the field was evaluated by challenge trial in which 60 per cent protection was obtained, or by monitoring the incidence of Newcastle disease in vaccinated and unvaccinated birds. There was no report of Newcastle disease outbreaks in the vaccinated birds during the two-year period of the field trial. The ease in administering the food pellet vaccine makes it readily accepted by the farmers.
This paper reports the experimental transmission of a bird parasite into jirds. Infective larvae of Cardiofilaria nilesi obtained from laboratory colonized Coquillettidia crassipes mosquitoes which had fed on an infected chicken were inoculated subcutaneously into jirds. The number of larvae per jird varied from 10 to 228. Microfilaraemia appeared 22 to 89 days after inoculation of the infective larvae. Experiments were carried out with 24 jirds through six generations extending over a period of 22 months and 17 produced patent infections. At present 8 infected jirds are being maintained in the laboratory; their patent periods ranging from 6 to 13 months. However, the longest patent period observed was about thirteen months. The percentage of adults recovered in autopsied jirds ranged from 0 to 40 with an average of 16. The chicken showed a microfilarial periodicity with the peak microfilarial density around 2200 hours. However, in jirds there was a change in sub-periodicity. This model in the jird may be very useful for the screening of filaricides and in immunological work.
A case of Newcastle disease virus infection in a female laboratory technician is reported for the first time in Malaysia. Infection was acquired by droplet infection of the eye while grinding infected chicken in the laboratory. The case was confirmed by isolation of Newcastle disease virus from an eye swab taken from the subject on the first day of clinical signs. A four-fold rise of haemagglutination-inhibition titre was shown when sera on the third day of infection and 15 days later were compared.
An experiment was carried out to estimate the meat quality characteristics of village chickens (Gallus gallus) fed diets supplemented with dry leaves of Borreria latifolia (BL) used as a potential antioxidant source in chicken feed. In this study, 252 sexed 9-week-old village chickens with mean live body weight of 1,525.4 g for males and 1,254.1 g for females were divided into 7 groups (each group 18 birds) for each sex represented in 2 experiments. The first experiment was to evaluate the antioxidant activity of BL and the effect on meat quality through a comparison with Rosmarinus officinalis (RO); hence, 3 groups were conducted and included: T1 (control), basal diet without supplementation; T2, basal diet with 1% of BL; T3, basal diet with 1% of RO. T2 and T3 significantly affect pH value, lipid oxidation, cooking loss, and overall acceptability compared to T1, while no significant difference was observed between the dietary groups in respect of drip loss, color, tenderness, fatty acid profile, and meat composition. Furthermore, a significant effect of sex on lipid oxidation, pH, yellowness, and fatty acid profile was observed. There was no significant effect of sex on WHC, tenderness, lightness, redness, and sensory evaluation. A significant influence of postmortem aging period was detected on lipid oxidation, pH, tenderness, cooking loss, and redness. The obtained result in this study revealed a significance in the interaction of herb by sex in pH parameter and between herb and sex, herb by aging period, sex by aging period, and the herb by sex by aging period interactions with regard to lipid oxidation test. The second experiment was to estimate the effect of 3 different levels of BL on meat quality. Four groups were provided and involved: T1 (control), basal diet without supplementation; T2, basal diet with 1.5% of BL; T3, basal diet with 2% of BL; and T4, basal diet with 2.5% of BL. The result of this study showed a significant effect (P