Monitoring of Newcastle Disease Virus Vaccine Strain Replication in Embryonated Chicken Eggs by Reverse Transcription-Polymerase Chain Reaction.

The knowledge of virus and replication kinetics plays a key role in developing a vaccine. This study aimed to monitor the replication process and determine the best harvesting time of the Newcastle disease virus (NDV) V4 vaccine strain in the allantoic fluids of specific pathogen-free (SPF)-embryonated chicken eggs (ECEs) by reverse transcription-polymerase chain reaction (RT-PCR), hemagglutination (HA), and egg infective dose 50% (EID50) tests. For this purpose, the V4 vaccine strain of the virus was intra-allantoically inoculated into 96 10-day-old SPF-ECEs at the rate of 0.1 mL/ECE. The allantoic fluids of the inoculated eggs were collected from six eggs at six-hour intervals up to 96 hours post-infection (hpi). The harvested suspensions were confirmed to contain the NDV by the mentioned serologic and molecular techniques. Based on the results, the virus was first detected at 36 hpi in ECEs by RT-PCR. The peak of HA and EID50 titers in allantoic fluids started at 42 hpi, and the titers were at the highest level until the end of the experiment. The results indicated that the best virus harvesting time for the NDV V4 vaccine strain in ECEs is between 42-60 hpi. These findings pave the way for adequate and enhanced production rate, immunogenicity, and cost-related parameters in the V4 Newcastle vaccine development.

Effects of Selected Adjuvants on Immunogenicity and Protectivity of Pasteurella multocida Bacterin Vaccine in Chickens.

Avian pasteurellosis (fowl cholera) is an important disease affecting domestic and wild birds all over the world. Although the capsular type A of Pasteurella multocida is mostly involved, other capsular types are occasionally incriminated. The present study aimed at investigating the effect of some adjuvants on immunogenicity and protectivity of P. multocida bacterin in chickens, compared to an Iranian commercial vaccine. Eight-week-old chicken pullets were double vaccinated with an interval of three weeks. Vaccine immunogenicity testing was conducted using an in-house indirect enzyme-linked immunosorbent assay and assessing serum antibody titers at 7, 14, and 21 days post-primary and 14 days post-secondary immunization. The possible adverse effects were recorded by a poultry-disease expert. For evaluating the vaccine protection rate, chickens were subjected to 2×Lethal Dose 50%of a virulent P. multocida strain two weeks post-secondary immunization. The rate of live and normal animals was regarded as protection rate 7days after the exposure. The findings showed that oil adjuvants Montanide ISA 70-and Montanide ISA 71-containingvaccines (with or without saponin) caused a powerful immune reaction than the aluminum adjuvanted vaccine and commercial vaccine (P<0.05). Significant protection against challenge was merely induced by the oil adjuvanted vaccines (P<0.05). The majority of the studied chickens showed inflammation at the injection site (yellow) throughout the trial. Vaccines made by Montanide ISA 70 and Montanide ISA 71 are novel and effective inactivated vaccines that are able to cause significant protection to fowl cholera disease.

Restriction Fragment Length Polymorphism Typing of Staphylococcus aureus Strains Isolated from Bovine Mastitis and Dairy Products in Ahvaz, Iran, Using of Digested Coagulase Gene.

Staphylococcus aureus is a major pathogen in the transmission of diseases from animals to humans and vice-versa.Various infections, such as mastitis in cattle, sheep and goats, as well as gastroenteritis due to food poisoning in humans are the most frequent problems caused by S. aureus. The bacteria also lead to severe economic losses in dairy industry. A major virulence factor for the organism is encoded by the coagulase (coa) gene. This study aimed to assess the polymorphisms of the coa gene in S. aureus strains isolated from bovine mastitis and dairy product samples in Ahvaz, Iran. The results showed that out of 91 S. aureus, 80 (87.91%) isolates were positive for coa gene(s). In total, nine different polymerase chain reaction (PCR) products were obtained for coa-positive isolates. A single band was detected in coa PCR with a size ranges from 370 to 830 bp in most isolates (n=77, 96.25%). For three isolates (3.75%), two amplification products were obtained. A PCR product of an estimated size of 590 bp was most frequent, as obtained for 48 (60.00%) isolates. Whereas, 370 and 830 bp PCR products were the least presented, for two (2.50%) and one (1.25%) isolate, respectively. Subsequently, for restriction fragment length polymorphism (RFLP), typing of coa gene and AluI restriction enzyme were used for the digestion of the products. AluI for most of PCR products generated a unique pattern; however, four PCR products (the sizes ranged 750, 670, 590, and 510 bp) generated three or more patterns. Based on AluI RFLP of coa gene, the isolates were classified into 23 groups. Two groups of isolates were dominant, making 45% of the total. According to the findings, one or two types of coa RFLP were dominant among samples that were infected with more S. aureus isolates belonging to different coa RFLP types.

Detection of Mycoplasma gallisepticum and Mycoplasma synoviae among Commercial Poultry in Khouzestan Province, Iran.

Mycoplasmas are important avian pathogens, which can cause both respiratory disease and synovitis in poultry that result in considerable economic losses to the poultry industry all over the world. The aim of this study was to determine the prevalence of Mycoplasma gallisepticum and Mycoplasma synoviae infections among commercial poultry flocks in Khouzestan province, Iran, using the polymerase chain reaction (PCR) technique. Totally, 290 tracheal swab samples were collected from 19 broiler flocks and 4 layer-breeder flocks, with or without respiratory signs, in different areas of Khouzestan province within six months. The PCR tests were applied for the specific amplification of 16S rRNA (185 bp) and vlhA (392 bp) genes. Out of 100 swab samples obtained from the layer-breeder flocks, 1 and 72 specimens were positive for M. gallisepticum and M. synoviae, respectively. In this regard, out of the 4 layer-breeder flocks, 1 (25%) and 4 (100%) flocks were positive for M. gallisepticum and M. synoviae, respectively. However, none of the studied broiler flocks were M. gallisepticum- or M. synoviae-positive. According to the results, the PCR technique could be concluded as a rapid method for the accurate identification of M. gallisepticum and M. synoviae infections in commercial poultry flocks. The results were indicative of the low prevalence of M. gallisepticum in the studied flocks in Khouzestan province. On the other hand, M. synoviae was widely distributed among layer-breeder flocks in this province.