Effects of embryonic thermal manipulation on the immune response to post-hatch Escherichia coli challenge in broiler chicken.

Background and Aim: Thermal manipulation (TM), exposure to mild heat shock during embryogenesis, which is a critical developmental period of broiler chickens, improves tissue stability, oxidative stress response, and immune response during heat stress. Thermal manipulation could be more cost-effective than other methods to boost the immune response. This study aimed to evaluate the impact of TM during embryogenesis, concomitant with an Escherichia coli challenge, on body weight (BW), body temperature (Tb), and splenic mRNA expression of cytokines (Interleukin [IL]-1ß, IL-2, IL-6, IL-8, IL-12, IL-15, IL-16, IL-18, and interferon [IFN]-γ) in poultry. Materials and Methods: A total of 740 fertile eggs were procured from a certified Ross broiler breeder. The eggs were divided into two incubation groups: the control and TM groups. The eggs in the control group were kept at 37.8°C air temperature and 56% relative humidity (RH) during incubation; eggs of the TM group were incubated under standard conditions, except for embryonic days 10-18, during which they were incubated at 39°C and 65% RH for 18 h daily. On the 7th day of incubation, eggs with dead embryos were excluded. After hatching was complete, each group was further subdivided into saline-treated or E. coli-challenged groups. The E. coli (serotype 078 with the dose of 1.5 × 105 colony-forming unit/mL) challenge was performed when the birds were 20 days old. Body weight and Tb measurements were taken on post-hatch days 20, 21, 23, and 25. Splenic mRNA expression of cytokines (IL-1ß, IL-2, IL-6, IL-8, IL-12, IL-15, IL-16, IL-18, and IFN-γ) was analyzed by real-time quantitative polymerase chain reaction. Results: Following the E. coli challenge, the TM-treated group's body performance parameters (BW and Tb) were significantly increased compared with the control group. Body weight was higher in the TM group than in the control group (p < 0.05); Tb was lower in the TM group than in the control group (p < 0.05). The mRNA levels of IL and IFN-γ were more stable and moderately induced in the TM group compared with the control group. Thermal manipulation altered the basal mRNA levels of ILs and IFN-γ and changed their expression dynamics after the E. coli challenge. Conclusion: Thermal manipulation during embryogenesis could boost the immune system response to E. coli.

Longitudinal and abattoir-based surveillance of MERS-CoV in camels in Jordan, 2018-2020.

To generate baseline information to help better understand the antibody kinetics and nasal shedding dynamics of MERS-CoV in camels in Jordan, a longitudinal surveillance study was conducted in two phases; phase 1 was between December, 2018 and January, 2019 and phase 2 between August and December 2020. In each phase, two camel herds were studied. These herds were located in Al-azraq and in Al-ramtha area and were named Al-azraq and Al-ramtha herds, respectively. The same camel herd of Al-zarqa area was sampled in both phases while two different camel herds, one in each phase, were sampled in Al-ramtha area. Blood and nasal swabs were collected from same selected animals in all visits to each herd in both phases. Additionally, nasal swabs and retropharyngeal lymph node tissue samples were collected from sixty-one camels slaughtered at Al-ramtha abattoir during phase 2 to enhance virus isolation opportunities and phylogenetic analysis. All sampled animals from Al-azraq camel herd were either borderline or seropositive on spike 1 based ELISA assay and negative on quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) in both phases. In Al-ramtha camel herds, an unsteady pattern prevailed in animals' seropositivity in both phases and viral RNA was detected in all animals in the end of phase 1 and in one animal during phase 2. For the seroconversion, anti-MERS-CoV spike 1 antibodies were detected in two animals in phase 1 in the first collection only. While, in phase 2, intermittent seroconversion pattern was observed in several samples over time of collections that ended with all animals became seropositive in the last collection (after nineteen days from viral RNA detection). In addition, viral RNA was detected in nasal swabs of 3 slaughtered camels. Phylogenetic analysis of a partial fragment of spike 1 gene sequences of all MERS-CoV isolates clustered together with clade B of MERS-CoV. This cluster contains all MERS-CoV sequences obtained either from camels or human sources in the Arabian Peninsula indicating the continuous circulation of this clade also in Jordan.

First detection of foot-and-mouth disease virus O/ME-SA/ Ind2001e sublineage in Jordan.

Foot-and-mouth disease (FMD) is a highly contagious vesicular disease that is caused by the FMD virus (FMDV). This disease affects both wild and domestic cloven-hoofed animals, and the latter of which includes cattle, swine, sheep and goats. FMD is endemic to Jordan and has a severe impact on the productivity of domestic livestock. In January 2017, FMD outbreaks were detected in different animal species across Jordan, resulting in high mortality rates among young lamb and goat populations as well as causing classic FMD symptoms in cattle. In this study, clinical specimens were collected from animals affected by FMD. The results obtained from sequencing the VP1 gene place the studied FMDV isolate within the FMDV O/ME-SA/ Ind2001e sublineage. Phylogenetic analysis of VP1 suggests that the O/JOR/1/2017 isolate is very similar to that of viruses isolated from Saudi Arabia in 2016. The possible introduction of this strain to Jordan might occur through transboundary animal movement or other transmission routes from Saudi Arabia, a neighbouring country.

Genetic diversity and population structure of local and exotic sheep breeds in Jordan using microsatellites markers.

AIM: This study was conducted to study the genetic and population structure of local (Awassi) and exotic (Romanov, Charollais, Assaf, Awassi, and Suffolk) sheep breeds in Jordan using eight microsatellite markers. MATERIALS AND METHODS: A total of 125 sheep were used (25 from each breed) in the study. The number of alleles (A), the mean values of observed (Ho) and expected (He) heterozygosity, polymorphism information content (PIC), fixation index as a measure of heterozygote deficiency or excess, and Hardy-Weinberg equilibrium (HWE) were analyzed using PopGen and CERVUS softwares. Nei's standard genetic distances among breeds and dendrogram of unweighted pair group method with arithmetic mean (UPGMA) were calculated and constructed using PopGen software. RESULTS: A total of 40 alleles were detected with an average number of alleles of 5. The mean Ho value was higher than the mean He value for all breeds. Awassi breed showed the highest average PIC value while Romanov had the lowest. There was a significant (p<0.05) deviation from HWE at each locus within and between breeds. Deviations from HWE were found to be highly significant for all markers except OARFCP304 locus. The genetic distance estimates revealed a close relationship between Romanov and Charollais and between Awassi and Charollais. In the UPGMA dendrogram, Charollais, Romanov, and Awassi breeds were placed together in one main cluster while Assaf was in a different subcluster. Awassi was placed alone in a second main cluster. CONCLUSION: Results of this study offer insight toward the genetic conservation of the studied breeds and a base on which breeding plans can be made.

Mycoplasma bovis arthritis and pneumonia in calves in Jordan: An emerging disease.

AIM: Clinical, microbiological, molecular, and pathological assays were undertaken to characterize an outbreak of increasingly reported signs of unresponsive arthritis and pneumonia of Mycoplasma bovis infection in young calves in Jordan. MATERIALS AND METHODS: Clinical history of the affected bovine herd was investigated for the presence of respiratory and/or joint problems. Two calves with such history were clinically examined and necropsied. Representative tissues were sent for microbiological, molecular, and pathological examinations for M. bovis infection. RESULTS: The outbreak started in a herd of 220 nursing calves, 2 months before the receiving of two calves for postmortem examination. Clinically, respiratory signs and infection of one or more joints dominated in the affected calves. The morbidity and case fatality rates were 27.27% and 61.7%, respectively. The left carpal joint was markedly swollen in both calves and exhibited necrofibrinous to granulomatous arthritis in varying degrees of severity. The anteroventral lung lobes in both calves were consistently affected and revealed multifocal to coalescing severe necrogranulomatous and fibrinopurulent bronchopneumonia. Microbiological and molecular findings confirmed the pathological examination. Furthermore, bovine viral diarrhea (BVD) was diagnosed in one calf by histopathology and polymerase chain reaction. CONCLUSION: This investigation reports the first outbreak of M. bovis infection in calves located in Jordan that could occur concurrently with BVD.

Comparative analysis of virulence and resistance profiles of Salmonella Enteritidis isolates from poultry meat and foodborne outbreaks in northern Jordan.

This study was conducted to isolate Salmonella Enteritidis from poultry samples and compare their virulence and antibiotic resistance profiles to S. Enteritidis isolated from outbreaks in northern Jordan. Two hundred presumptive isolates were obtained from 302 raw poultry samples and were subjected to further analysis and confirmation. A phylogenic tree based on 16S rRNA sequencing was constructed and selected isolates representing each cluster were further studied for their virulence in normal adult Swiss white mice. The most virulent strains were isolated from poultry samples and had an LD 50 of 1.55 × 10 (5) CFU, while some of the outbreak isolates were avirulent in mice. Antibiotic resistance profiling revealed that the isolates were resistant to seven of eight antibiotics screened with each isolate resistant to multiple antibiotics (from two to six). Of the poultry isolates, 100%, 88.9%, 77.8%, 66.7%, and 50% showed resistance to nalidixic acid, ciprofloxacin, ampicillin, cephalothin, and cefoperazone, respectively. Two outbreak isolates were sensitive to all tested antibiotics, while 71.4% were resistant to cefoperazone and only 28.6% showed resistance to nalidixic acid. Salmonella outbreak isolates were genetically related to poultry isolates as inferred from the 16S rRNA sequencing, yet were phenotypically different. Although outbreak strains were similar to poultry isolates, when tested in the mouse model, some of the outbreak isolates were highly virulent while others were avirulent. This might be due to a variation in susceptibility of the mouse to different S. Enteritidis isolates.

Thermal manipulation during chicken embryogenesis results in enhanced Hsp70 gene expression and the acquisition of thermotolerance.

The aim of this study was to evaluate the effect of thermal manipulation (TM) during embryogenesis on hatchability, growth performance and thermotolerance acquisition parameters during thermal challenge (TC). Seven-hundred and fifty fertile chicken eggs were divided randomly into three groups (250 eggs each): control group was maintained at 37.8°C and 56% relative humidity (RH), TM1 was subjected to TM at 38.8°C for 6h and 65% RH during embryonic days (ED)10-18 and TM2 was subjected to TM at 38.8°C for 18 h and 65% RH during ED10-18. Hatched chicks from each treatment group were then randomly divided into two sub-treatment groups (Naive and TC). Chicks in TC groups were subjected to TC by adjusting room temperature to 41.0°C for 6h on days 3, 7, and 42 of age while naïve chicks were kept under regular conditions (25 ± 1°C and 50-60% RH). Percentage of hatched eggs was recorded and post-hatch chick performance was evaluated by recording chick body weight (BW). Chick's response to TC was evaluated by determination of body temperature (T(b)), plasma T3 and T4 levels, and muscle mRNA levels of Hsp70. There was a significant increase in muscle mRNA levels of Hsp70 during embryogenesis and during TC in post-hatch chicks. While hatchability was not adversely affected, the body weight in TM2 chicks was significantly higher at the end of the study period (42 days). Results of this study indicated a long-term enhancement of Hsp70 gene expression associated with improved thermotolerance acquisition in treated chicks without adversely affecting performance.

Detection and molecular characterization of bovine leukemia viruses from Jordan.

Bovine leukemia virus (BLV) is distributed worldwide. BLV has many effects on the health status and productivity of infected animals and is a potential risk for humans. In this study, we aimed to investigate the presence of and genotype bovine leukemia viruses on Jordanian dairy farms. Nested PCR coupled with RFLP and direct sequencing of a partial fragment of the env gene were carried out. Two BLV genotypes were found, genotypes 1 and 6. These genotypes were identified by nested PCR-RFLP of 444 bp of the env gene by restriction digestion with HaeIII, Bcl I and Pvu II. However, BLV-Jordan-10 seems to represent an entirely new genotype in our phylogenetic analysis. The nucleotide sequence identity between these two Jordanian BLV genotypes (1 and 6) was 96.2 %. The nucleotide sequence identity between Jordanian BLV genotype 1 and other reference BLV genotype 1 strains ranged from 99 % to 99.5 %. The nucleotide sequence similarity of the Jordanian BLV genotype 6 to other BLV genotypes ranged from 90 % to 96.7 %. A neutralizing motif and CD8(+) T-cell epitope were found in the env protein of both Jordanian isolates. In this study, we documented the presence of two BLV genotypes (1 and 6) on Jordanian dairy farms.

Prevalence and risk factors associated with Chlamydophila abortus infection in dairy herds in Jordan.

A cross-sectional study was carried out to determine seroprevalence and to identify risk factors associated with Chlamydophila abortus infection in 62 nonvaccinated dairy herds (671 cows) in Jordan between January and June 2007. Information regarding herd management was recorded through a personal interview with farmers. Antibodies against C. abortus were detected using an ELISA test kit. Chi-square analysis and multivariable logistic regression model were used to identify risk factors associated with C. abortus seropositivity. The true prevalence of antibodies against C. abortus in individual cows and cattle herds were 19.9 % and 66.3 %, respectively. Univariable Chi-square analysis revealed three variables with P ≤ 0.25 that were further offered to multivariable logistic regression analysis. Small-sized herds were identified as a risk factor for seropositivity to C. abortus, while sweeping followed by water hosing and using disinfectants were identified as protective factors. Cows in the age groups of >8 and ≤ 10 years old and >2 and ≤ 6 years old had the highest and lowest significant seroprevalence to C. abortus, respectively. Results of this study indicated that C. abortus is highly prevalent in Jordan's dairy herds and Chlamydophila infection could be controlled by applying strict biosecurity measures in the dairy farms.

Molecular characterization of a recent Newcastle disease virus outbreak in Jordan.

Newcastle disease (ND) is a highly contagious viral disease and is a continuous threat to the poultry industry worldwide. In the early months of 2011, several devastating ND outbreaks occurred in Jordan affecting broilers, layers and breeders. The fusion gene of the isolated Newcastle disease virus (NDV) was partially amplified by RT-PCR, then directly sequenced. The NDV isolates were found to have the motif112RRQKRF117. This motif and a mean death time (MDT) of 46 h are indicative of the velogenic nature of these NDV isolates. Phylogenetic analysis showed that the new NDV strain belongs to the lineage 5d (Aldous et al., 2003) and is closely related to the Chinese strain SG/Liaoning/2009. NDV outbreaks in 2010 and 2011 have been noted in neighboring countries. Based on the high nucleotide similarity between our isolated NDV isolates and the Chinese NDV strain, the origin of these recent NDV isolates might be from China.

Female reproductive tract anatomy of the endangered Arabian oryx (Oryx leucoryx) in Jordan.

Female reproductive anatomy of the Arabian oryx is unknown. In this study, reproductive tracts of seven female Arabian oryx (aged 2 to 7 years) were examined to characterize their reproductive anatomy. Observations and measurements were obtained in situ from dead animals during necropsy. Animals were allocated into two groups: cycling (n = 3; follicles or corpora lutea present) and not-cycling (n = 4; follicles or corpora lutea absent). Different reproductive tract segments for each animal in both groups were measured using a digital caliper. The mean, SD and range for each reproductive tract segment were generated and compared between groups. Female oryx reproductive anatomy share some anatomical characteristics with that of domestic ruminants except that the oryx uterus has no distinct uterine body and the cervix has two internal openings for each respective uterine horn. In addition, there were more than 8 rows of caruncles within each uterine horn. There were significant differences in the length and width (P < 0.05), but not in height, of both the right and left ovaries between cycling and not-cycling animals (P > 0.05). Posterior and anterior vaginal lengths varied between cycling and not-cycling groups (P < 0.05). Length of right and left oviducts, left and right uterine horns, cervix and vulva did not vary between cycling and not-cycling groups (P > 0.05). Defining this unique morphology of female Arabian oryx reproductive anatomy will help in the development of appropriate reproductive techniques in order to propagate this endangered species and control its reproduction.

Ovine and caprine toxoplasmosis (Toxoplasma gondii) in aborted animals in Jordanian goat and sheep flocks.

Two hundred and fifty-five biological samples (106 aborted foetal tissue samples and 149 blood samples from aborted sheep and goats) were collected from 188 animals during the lambing season from September 2009 to April 2010 from the Mafraq region of Jordan. The sampled animals belonged to 93 goat and sheep flocks that had cases of abortion. A total of 169 (66.3%) biological samples were collected from sheep and 86 (33.7%) from goats. Seventy-six (29.8%) biological samples (45 blood and 31 tissue samples) were positive for Toxoplasma gondii by PCR assay. The positive samples were obtained from 43 sheep and 23 goats. The overall toxoplasma-specific prevalence rate was 35.1% (66/188). Forty flocks (43%) had at least one T. gondii PCR-positive animal. The risk factors related to flock health status and farm management that are hypothesized to be associated with T. gondii PCR positivity were also assessed using multiple logistic regressions. The presence of cats (OR = 4.74), a large flock size (OR = 2.76) and the method of disposing the aborted foetuses (OR = 3.77) were all statistically significant (P<0.05) risk factors that were positively associated with toxoplasma positivity in goat and sheep flocks.

Seroprevalence and risk factors for bovine brucellosis in Jordan.

We investigated the seroprevalence and risk factors for Brucella seropositivity in cattle in Jordan. The sera from 671 cows were randomly collected from 62 herds. The antibodies against Brucella were detected using a Rose Bengal plate test and indirect ELISA. A structured questionnaire was used to collect information on the cattle herds' health and management. A multiple logistic regression model was constructed to identify the risk factors for Brucella seropositivity. The true prevalence of antibodies against Brucella in individual cows and cattle herds was 6.5% and 23%, respectively. The seroprevalence of brucellosis in cows older than 4 years of age was significantly higher than that in the younger cows. The seroprevalence of brucellosis in cows located in the Mafraq, Zarqa and Ma'an governorates was significantly higher than that of the other studied governorates. The multiple logistic regression model revealed that a larger herd size (odd ratio OR = 1.3; 95% CI: 1.1, 2.6) and mixed farming (OR = 2.0; 95% CI: 1.7, 3.7) were risk factors for cattle seropositivity to Brucella antigens. On the other hand, the use of disinfectants (OR = 1.9; 95% CI: 1.1, 2.1) and the presence of adequate veterinary services (OR = 1.6; 95% CI: 1.2, 3.2) were identified as protective factors.