Alternative therapeutic approach of ovine subclinical mastitis using the ethanolic roots extract of Capparis spinosa.

Background: Over the past decades, Capparis spinosa has been considered a traditional therapy for relieving different illnesses. Mastitis causes a decrease in milk production and is usually treated with injectable and intra-mammary antibiotics. Aim: Investigating the therapeutic effects of C. spinosa root extract on subclinically mastitic ewes. Methods: Totally, 164 lactating ewes were selected randomly from the flocks that existed in some areas belonging to Al-Najaf City (Najaf, Iraq) from September to December (2022). Each study animal was subjected to direct sampling of milk before and once each week for 6 weeks (42 days) post treatment to be tested directly by the California mastitis test (CMT). Results: Concerning phytochemical testing of ethanolic root extract, the findings revealed a significant increase in the concentration of alkaloids, flavonoids, polyphenols, and tannins when compared to other components such as coumarins, saponin, glycosides, amino acids, and steroids. In this study, there were 44.51% infected ewes with subclinical mastitis, involving 25.61%, 13.41%, and 5.49% for scores 1, 2, and 3, respectively. In comparison with pre-treatment week, insignificant alteration was seen in the values of all scores in therapeutic week 1. However, significant differences were initiated in values of score 0 in week 2; score 0 and score 2 in week 3; score 0, score 1, and score 2 in week 4; and values of all scores in weeks 5 and 6. Conclusion: This represents the first Iraqi study aimed at the treatment of subclinical mastitis in sheep using the root extract of C. spinosa. Phytochemical testing of ethanolic extract revealed the presence of variable amounts of chemical compounds that reflect their effects on treated animals by decreasing the number of infected ewes with the disease. Moreover, studies are greatly important to estimate the therapeutic effects of other parts of C. spinosa such as leaves and seeds, on the disease and other animal diseases.

Serological identification of MERS-CoV in camels of Wasit province, Iraq.

Background: Since the first human case of Middle East Respiratory Syndrome (MERS) caused by Coronavirus (MERS-CoV) in 2012, several evidence bases have shown one-humped camels as the main reservoir host, from which infection is transmitted to humans. Aim: Serological investigation of MERS in dromedary camels in Wasit province (Iraq), detection severity of infection, and association to some risk factors. Methods: A total of 455 dromedary camels were selected randomly from two main districts in Wasit province, Iraq, during January and April (2023). Sera of all study camels were examined by enzyme-linked immunosorbent assay (ELISA), and titers of positive study animals were categorized according to their severity. Results: Serological testing yielded 37.58% positive animals for MERS infection. According to the severity of positive ODs (titer), a total of 53.22%, 30.99%, 12.28%, and 3.51% showed mild, moderate, strong, and very strong infections, respectively. Regarding risk factors, significant elevation in seropositivity was seen in camels of >3-6 and >6 years old and reduced in camels of £3 years old with an elevated risk of MERS with increased age. Regionally, seropositivity and relative risk were increased in the camels of Shaykh Sa'd when compared with Al-Numaniyah. Regarding sex, no significant variation was detected between seropositive females and males; however, male camels appeared at higher risk than females. Association between the severity of MERS infection and risk factors revealed that there was a significant increase in mild and moderate infections in female camels of >6 years old; whereas strong and very strong infections were seen in male camels of 33-6 years old. Mild and very strong infections were recorded in Shaykh Sa'd; while moderate and strong infections in Al-Numaniyah. Conclusion: The study indicated a longstanding existence of MERS-CoV in camels of Wasit province; therefore, recent infections or active viral excretion are required for confirmation by molecular approaches.

Role of nuc gene in Staphylococcus aureus to phagocytic activity in different cattle infections.

Background: Staphylococcus aureus is a typical pathogenic agent that causes several morbidities and mortalities which variable largely following the severity of bacteria and activity of host immunity. Aim: Isolation of S. aureus from different cattle infections, molecular detection of nuc gene in positive S. aureus isolates, and identification of the effectiveness of the phagocytic activity. Methods: Totally, 100 cattle with various infections (25 wounds, 25 abscesses, 25 nasal discharges, and 25 ear swaps) were selected and subjected to collection of swabs under controlled conditions. All collected samples were cultured on mannitol salt agar (MSA) and assessed by biochemical tests. Targeting the nuc gene, all study MSA positive isolates were examined molecularly by conventional polymerase chain reaction (PCR), and then subjected to antibiotic susceptibility test. Jugular venous blood was collected from all infected animals in addition to 20 healthy cattle that were selected as a control group to estimate the phagocytic activity of S. aureus isolates. Results: The findings of MSA culture revealed a total of 80 positive samples of S. aureus as 23, 21, 20, and 16 positive isolates for nasal discharge, abscess, wound, and ear swab, respectively; based on its morphology, cultural trait, and biochemical test. Subsequently, PCR assaying of MSA-positive isolates demonstrated an overall 59 positive samples as 14, 16, 12, and 17 positive isolates for nasal discharge, abscess, wound, and ear swabs, respectively. Antibiotic susceptibility testing of S. aureus-positive PCR isolates reported a significantly high sensitivity to chloramphenicol and vancomycin, and a high resistance to penicillin. Finally, there was a considerable decline in phagocytic activity in particular 2 weeks post-infection as a result of bacterial invasion. Conclusion: This study shows a high prevalence of S. aureus in cattle infections, and the protocol includes a regular screening of cattle infection and suitable therapy based on antibacterial susceptibility test is of great importance in the long-term control of the pathogen. However, additional molecular studies targeting other genes of S. aureus and the role of immune markers in different infections should be aimed.

Molecular prevalence and phylogenetic confirmation of bovine trichomoniasis in aborted cows in Iraq.

Background and Aim: Bovine trichomoniasis, caused by Tritrichomonas foetus, is a venereal disease that is distributed in many countries, including Iraq. Compared with other abortive infectious diseases, prevalence of T. foetus is expected to be relatively low in the field by veterinarians. This study aimed to estimate the prevalence of T. foetus in aborted cows by conventional polymerase chain reaction (PCR) and phylogenetic analysis of local T. foetus isolates was documented in the National Center for Biotechnology Information as the first sequenced isolates from Iraq. Materials and Methods: Vaginal fluids were collected from 62 aborted cows and examined by PCR. Data were reported for the following parameters: Vital signs (body temperature and respiratory and pulse rates), age (<4, 4-8, and >8 years), reproductive health status (premature calving, embryonic death, pyometra, and healthy newborn), breed (pure or crossbred), type of breeding (natural or artificial), bull-to-cow ratio (1:<10, 1:10-20, and 1:>20), contact of cow with bull(s) from other farmers (yes or no), and contact with stray animals (dogs and cats). Results: A total of 20.97% of aborted cows were positive for T. foetus. Phylogenetic analysis for 10 positive local T. foetus isolates demonstrated high identity with the Thai (MN560972.2) and Chinese (MH115435.1) isolates, with an identity range of 98.8%-99.5% and 98.6%-99.3%, respectively. Clinical data showed that the vital signs differed insignificantly between cows positive and negative for T. foetus. Prevalence and risk of infection increased significantly in <4-year-old, early calving, embryonic death, crossbred, and naturally inseminated cows that had direct contact with bulls from other farmers, and contact with stray animals. Fetal pneumonia and death of premature calves were significant among positive aborted fetuses. Conclusion: Tritrichomonas foetus is highly prevalent in aborted cows in Iraq and phylogenetic analysis demonstrated an identity between the local and global isolates, that is, Thai and Chinese, of cats.

Effectiveness of raw bacteriocin produced from lactic acid bacteria on biofilm of methicillin-resistant Staphylococcus aureus.

Background and Aim: Probiotics are proven beneficial to health since they enhance immunity against dangerous pathogens and increase resistance to illness. Bacteriocin produced by lactic acid bacteria (LAB), demonstrates a broad inhibitory spectrum and therapeutic potential. This study aimed to isolate LAB-producing bacteriocin and investigate the effect of crude bacteriocin on biofilm from methicillin-resistant Staphylococcus aureus (MRSA). Materials and Methods: This study used randomly collected 80 white soft local cheeses (40 each from cows and sheep) from different supermarkets in Basrah Province. The obtained samples were cultured and the bacterial suspension of S. aureus was prepared at 1.5 × 108 cells/mL. The crude bacteriocin extracted from LAB was obtained, and the tube was dried and inverted to detect the biofilm loss at the bottom. Results: There were 67 (83.75%) LAB isolates. Among 40 milk samples collected directly and indirectly, there were 36 (83.33%). Staphylococcus aureus isolates based on conventional bacteriological analysis and biochemical tests. Molecular testing was conducted to identify LAB and MRSA. Depending on genotypic results, the effect of white soft local cheese (cows and sheep) and the amplification results of the 16S rRNA gene were detected in 46 LAB isolates from white soft local cheese from cows and sheep. Based on the molecular identification of the mecA, results on Staphylococcus determined that only 2 of 36 isolates of S. aureus carried the mecA. Moreover, there were 26 (86.66%) isolates (MRSA) from samples of raw milk from local markets and subclinical mastitis in cows. The ability of LAB isolates was tested. The effects of bacteriocin production on preventing biofilm growth and formation were investigated. Results demonstrated that bacteriocin has high activity. Microtiter plates applied to investigate the ability of S. aureus to produce biofilms revealed that all isolates were either weak or moderate biofilm producers, with neither non-biofilm nor strong biofilm producers found among the tested isolates. Conclusion: Lactic acid bacteria demonstrate a high ability to produce bacteriocin. Crude bacteriocin from LAB has a restrictive effect on biofilms produced by MRSA; thus, it can be used to reduce the pathogenicity of this bacterium.

Molecular genotyping, histopathological and immunohistochemical studies of bovine papillomatosis.

Background: Bovine papillomatosis (BP) is considered the most common health problem in large cattle farms. Aim: This study attempts to confirm clinically suspected BP in cattle by polymerase chain reaction (PCR) assay, histopathology, immunohistochemistry (IHC), and genotyping analysis of local isolates. Methods: According to morphological appearance and lesion features, a cross sectional study of 54 clinically diagnosed BP cattle was assigned to this current investigation from May to August (2021) in Al-Kut district (Wasit Province, Iraq) private veterinary clinics using purposive sampling technique based on set criteria. The cattle were diagnosed clinically, and the tissues were collected and some fixed in 10% neutral buffered formalin and other stored frozen and examined by histopathological technique, IHC, and PCR assays. Results: Using PCR assay, all cattle were positive for the BPV L1 gene. According to detect the L1 gene, analysis of the phylogenetic tree showed that local BPV cattle isolates were closely related to the NCBI-BLAST BPV type-1 and type-2 of the Polish equine isolate (KF284133.1) and BPV Brazilian Bostaurus isolate (MH187961.1), respectively. Histological detection showed there were acanthosis, hyperkeratosis, epidermal thickening, severe infiltration of mononuclear cells, massive hemorrhage, dermal fibroplasias, multifocal spongiosis, moderate neovascularization, moderate to severe elongation of the retention ridge towards the dermis, parakeratosis, rings of calcification, and necrosis with nuclear pyknosis of some spinosum cells. Immunohistochemical findings of tumor necrosis factor-alpha, epidermal growth factor receptor and Fascin showed a significant variation in values of immunoreaction in the dermis and epidermis. These results ranged from negative (0) to mild positive (+1) to moderate positive (+2) reactions. Conclusion: The study provided essential molecular and genotyping data to improve our knowledge by emphasizing the crucial of IHC as an elegant diagnostic method to detect cellular alterations.

The role of endoplasmic reticulum stress in endometriosis.

Endometriosis is a chronic gynecologic disorder characterized by abnormal growth of endometrium-like tissues in the ectopic regions of the pelvic peritoneum. The pathophysiology of endometriosis is not completely understood; however, excessive endometrial cell proliferation together with resistance to apoptosis facilitates the migration, implantation, and survival of endometrial cells in the distant sites. Endoplasmic reticulum (ER) stress response (also called unfolded protein response) is a cellular defense mechanism triggered by ER stress. When severe enough, the so-called response initiates cell suicide, i.e., apoptosis. Therefore, therapeutic induction of ER stress in endometriotic cells could promote apoptosis and contribute to the management of disease. In this review, we discuss the pathogenic role of ER stress in endometriosis and the most recent findings regarding the induction of ER stress in connection with endometriosis.

The first study on confirmation and risk factors of acute and chronic canine distemper in stray dogs in Wasit Province, Iraq, using enzyme-linked immunosorbent assay and reverse transcription-polymerase chain reaction.

Background and Aim: In Iraq, stray dogs represent a critical population of free-roaming animals, which probably play a role in the transmission of different infections to other animals. Canine distemper is one of the most growing viral threats to carnivores in many countries worldwide, including Iraq. Therefore, this study was aimed to diagnose the disease using serological and molecular assay and the role of risk factors in the spreading infection. Materials and Methods: In all, 158 venous blood samples were collected randomly from stray dogs in rural and sub-urban areas of Iraq from May 2019 to December 2020. The samples were examined serologically using two enzyme-linked immunosorbent assay (ELISA) kits, immunoglobulin G (IgG) and immunoglobulin M (IgM), and molecularly by reverse transcription-polymerase chain reaction (RT-PCR) to detect and confirm chronic and acute infections. To determine the association between infection and various risk factors, the study animals were divided according to their locations, sexes, and ages. The age groups were ≤8 months (puppy), 1-3 years (young), and ≥3 years (old/mature). Results: ELISA result shows that 6.96% and 19.62% of dogs were seropositive for acute and chronic distemper, respectively. The titer of chronic infections (0.421±0.027) was significantly higher (p≤0.025) than that of acute canine distemper (0.337±0.016). On RT-PCR, 8.86% of dogs were found positive for distemper. Using RT-PCR as the gold standard, the sensitivity and specificity of the IgM ELISA kit were 75% and 98.63%, respectively, whereas the positive and negative predictivity were 81.82% and 97.96%, respectively. A significant variation (p<0.05) was observed in the distribution of positive findings among the different epidemiological risk factors. Compared with rural areas, positivity was significantly higher (p<0.05) in sub-urban areas on IgM (26.92%) and IgG (64.15%) ELISA and RT-PCR (34.62%). On IgM ELISA and RT-PCR, no significant differences (p>0.05) were found among the three age groups; however, positivity was significantly higher (p≤0.048) in the ≥3 years group (22.73%) on IgG ELISA. Furthermore, only IgG ELISA showed a significantly higher (p≤0.032) positivity rate in female dogs (25.23%) than in male dogs (7.84%). Conclusion: To the best of our knowledge, this is the first Iraqi study to demonstrate acute and chronic canine distemper in stray dogs, suggesting that the prolonged shedding of virus from positive dogs is a critical point in the epidemiology of the disease. Additional studies in dogs or other carnivores are required to establish baseline data on the prevalence of the disease in Iraq.

First molecular confirmation of Coenurus cerebralis in sheep and goats with neurological behaviors in Iraq.

BACKGROUND AND AIM: Coenurus cerebralis is the intermediate stage of the canine cestode, which infect sheep and goats, resulting mainly in neurological signs and causing direct and indirect economic losses. This study aimed to demonstrate the existence of C. cerebralis and to elucidate the role of this parasite in inducing neurological behaviors in sheep and goats. MATERIALS AND METHODS: On the basis of historical data on neurological signs, we subjected 76 animals (49 sheep and 27 goats) of different ages, sexes, and geographical areas for molecular examination of their blood samples using the polymerase chain reaction assay. RESULTS: Of the 76 animals, 23.68% tested positive for C. cerebralis infection. We found significant increases in infection (p<0.05) in sheep (26.53%) more than in goats (18.52%). Circling movement was prevalent significantly among both C. cerebralis-positive sheep and goats. The Nineveh region had a significant (p<0.05) increase in positive sheep and goats, and the sheep of all study regions were infected significantly (p<0.05) more than were the goats. We found no significant (p>0.05) variation between sheep ages ≥1-3 years and ≥3 years; however, both groups had a significantly (p≤0.043) higher positivity rate than did sheep ages <1 year. The findings of sheep ages <1 year and ≥1-3 years were significantly higher than those of the goats, but not for goats ages ≥3 years. Female sheep and goats showed a significant increase in positivity versus that for the males. CONCLUSION: To the best of our knowledge, this study is the first report in Iraq targeting detection of C. cerebralis in sheep and goats with neurological behaviors; therefore, additional studies involving different animals in other regions using molecular techniques are needed.

Molecular and histopathological identification of ovine neosporosis (Neospora caninum) in aborted ewes in Iraq.

AIM: The objective of the present study was to detect Neospora caninum DNA in the placenta of sheep and evaluate the association of risk factors to polymerase chain reaction (PCR) positive and histopathological analysis of the placenta and fetal tissue samples of aborted fetuses. MATERIALS AND METHODS: Fresh placenta from 51 aborted ewes was collected for PCR assay. Placental and fetal tissues of aborted fetuses, including brain, heart, liver, lung, and thymus, were collected for histopathological analysis, besides the risk factor data were obtained during the time of sampling. RESULTS: From 51 placentas examined by PCR, 13.73% appeared positive to N. caninum DNA. The relationship between PCR positive and the risk factors revealed a significant difference (p<0.05) in age of the dam, fetal age, feed source, water source, and the presence of other animals at farm, whereas the type of birth, stillbirth, and size of flock showed insignificant difference (p>0.05). Histopathological investigation of placental and fetal tissues of positive samples showed tissue cyst-like structure, necrotic foci, and infiltration of mononuclear cells. Other lesions were thickening in chorionic plate in placenta, severe vacuolization and death of neurons, microgliosis, demyelination, edema, and proliferation of astrocytes in brain. In addition, fibrous and fat deposition with stenosis in the heart, parenchymal necrosis, severe atrophy, vacuolization and hyalinization of hepatocytes, megakaryocyte, portal fibrosis in the liver, and interlobular septal thickening in lung without obvious lesions is seen in the thymus tissue samples. CONCLUSION: This is a unique study that confirmed N. caninum DNA in the placenta of aborted ewes in Iraq using PCR assay. Histopathological analysis of some aborted fetuses organs could provide a more confirmatory and reliable data for a significant role of neosporosis in increasing the rate of abortion in sheep, while the clinical data of risk factors could be used to control the transmission of N. caninum infection.

Molecular and histopathological confirmation of clinically diagnosed lumpy skin disease in cattle, Baghdad Province of Iraq.

AIM: This study aimed to confirm the clinically diagnosed cattle with lumpy skin disease (LSD) at Baghdad Province/Iraq from October 2018 to March 2019. MATERIALS AND METHODS: Molecular polymerase chain reaction (PCR) and histopathology were applied for the detection of LSD among 71 infected cattle issued for slaughter. RESULTS: Pre-slaughter clinical examination showed significant increases (p<0.05) in values of temperature (39.7±0.74°C), pulse (96.42±3.51), and respiratory (33.54±0.63) rates. Enlargement of lymph nodes (prescapular, supramammary, and prefemoral), lacrimation, mucopurulent nasal discharge, salivation, edema in limbs and head among severe infected cases, and marked fall in milk production was seen. An association of LSD to risk factors (age, gender, and areas) showed that there is significant elevation in prevalence of disease in >2-5 years (54.93%) rather than other age groups (>5 and <2 years)in females (73.24%) than males (26.76%); and in sub-rural (42.25%) and rural (39.44%) compared to urban (18.31%) areas. Postmortem examination appeared nodular lesions in upper parts of the digestive system (9.86%), rumen (2.82%), upper respiratory tracts (7.04%), and lung (4.23%). The PCR examination of P32 and thymidine kinase antigenic genes showed 90.14% and 60.56% positive samples, respectively. Histopathological analysis of nodular skin biopsies showed edema, hyperemia, acanthosis, severe hydropic degeneration, and hyperkeratosis in epidermis; whereas, mononuclear cell infiltration, inclusion bodies, and vasculitis seen in the dermis. CONCLUSION: PCR and histopathology assay could be a potential method to confirm the LSD infection concomitant with clinical examination.