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    The Association of Intrauterine Antimicrobial Resistant Trueperella pyogenes with Cytological Endometrial Inflammation Status in Repeat Breeder Cows
    (Zoological Soc Pakistan, 2022) Yazlik, Murat Onur; Mustak, Inci Basak; Unal, Gultekin; Kaya, Ufuk
    This study describes the presence of Trueperella pyogenes from routine bacteriological examination, sequence analysis for antimicrobial resistance of the identified bacteria, and the association of T. pyogenes with cytological endometrial inflammation status in 32 repeat breeder cow enrolled in the study. Intrauterine samples for bacteriological examination were collected with cotton swabs; samples for subclinical endometritis were collected with cytotape technique. Antimicrobial resistance to commonly used antibiotics was evaluated by disc diffusion method. Presence of the tetW gene responsible for tetracycline resistance was detected by PCR. Subclinical endometritis, breed, parity, calving history, early postpartum uterine infection, ovarian tissue findings, and body condition score were analyzed as potential factors for T. pyogenes presence in repeat breeder cows. A total of 15 isolates were identified as T. pyogenes. These were combined to analyze the results by both biochemical and sequence analysis of the 16s rDNA gene. Subclinical endometritis was detected in 37.5% cows. The T. pyogenes strains were highly resistant (100%) to bacitracin, tetracycline, neomycin, and oxytetracycline. All tetracycline resistant T. pyogenes strains were positive for the tetW gene. The variables had no effect on the presence of T. pyogenes in the repeat breeder cows. In conclusion, intrauterine T. pyogenes presence was not associated with subclinical endometritis, although T. pyogenes was detected in the later stages of lactation.
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    Expression patterns of inflammatory and oxidative stress-related genes in the uterine and ovarian tissues of dogs diagnosed with pyometra based on cervical patency status
    (Elsevier Science Inc, 2024) Ozkan, Huseyin; Yazlik, Murat Onur; Keceli, Hasan Huseyin; Vural, Sevil Atalay; Mustak, Inci Basak; Kaya, Ufuk; Altinbas, Yunus Furkan
    This study aimed to investigate the expression patterns of genes associated with inflammation and oxidative stress in ovarian and uterine tissues of dogs with pyometra, categorized by cervical status (open cervix or closed cervix), which influences disease severity. The control group comprised healthy animals undergoing elective ovariohysterectomy. Tissue inflammatory gene expression and Malondialdehyde (MDA) levels were determined while microbial and histopathological examinations were conducted, along with immunohistochemical evaluations. In the closed-cervix group, uterine TNF and IL6 were upregulated approximately 10- fold while IL10 was upregulated nearly 5-fold. TNF expression differed remarkably between the pyometra groups. In the closed-cervix group, PTGS2 and HMOX1 were upregulated approximately 5-fold whereas NFE2L2 expression was downregulated. The closed-cervix group also had the highest uterine MDA levels. Regarding ovarian tissue, MDA levels were higher in the closed-cervix group than in the open-cervix group while IL10 expression was lower in the closed-cervix group than the open-cervix group. In the closed-cervix group, NFE2L2 was downregulated whereas HMOX1 was upregulated. Uterine TNF levels were positively correlated with IL6, , IL10, , PTGS2, , and HMOX1, , but negatively correlated with NFE2L2. . IL6 was positively correlated with IL10, , PTGS2, , and HMOX1. . NFE2L2 was negatively correlated with IL6 and HMOX1. . IL10 was positively correlated with PTGS2 and HMOX1. . MDA was positively correlated with TNF, , IL6, , IL10, , PTGS2, , NFE2L2, , and HMOX1. . TNF levels were positively correlated with ovarian PTGS2, , and with IL6 and NFE2L2. . MDA was positively correlated with PTGS2 and HMOX1. . MDA could be an important biomarker for understanding the severity of pyometra. Moreover, TNF expression and its relationships with various studied parameters such as IL10 may contribute to treatment and prognostic biomarker studies in closed-cervix pyometra pathology.
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    The presence of virulence factor genes downregulates uterine AQP3 and alters glutathione peroxidase activity and uterine histopathology in canine pyometra
    (Wiley, 2024) Yazlik, Murat Onur; Mustak, Inci Basak; Ozkan, Hueseyin; Vural, Sevil Atalay; Kaya, Ufuk; Ozoner, Ozgur; Mutluer, Ipek
    Present study was designed to evaluate the role of virulence factor genes (papG, cnf1 and hylA) in the pathogenesis of canine pyometra. Antimicrobial susceptibility test and detection of virulence genes were performed Escherichia coli (E. coli) detected in uterine swab samples. Animals were divided into two groups based on the presence (VF+, n:14) or absence (VF-, n:7) of the virulence factor genes papG, cnf1 and hylA. Blood and tissue glutathione peroxidase activity, uterine histopathologic analysis and AQP3, ESR1, PGR, OXTR gene expressions were determined in both groups. Statistical analyses were performed using Stata version 15.1. All E. coli isolates were susceptible to amikacin, whereas resistant to ampicillin, amoxicillin/clavulanic acid and lincomycin. None of the isolates were susceptible to cefotaxime. E. coli isolates had at least one virulence gene. The most prevalent gene was fimH (100%), followed by fyuA (95.8%), usp (83.3%), sfa (75%), cnf1 and hlyA (70.8%) genes. Blood GPx activity was greater in VF+ animals. On the other hand, uterine tissue GPx activity was lower in VF+ group compared to the control group. Expression levels of AQP3 were upregulated more than fivefold in VF-dogs compared to the control group. In addition, AQP3 expression levels were found approximately threefold higher in VF (-) than VF (+) group (p < .05). Varying degree of inflammation noted for all animals with pyometra, but the presence of bacteria noted only in VF+ animals. In conclusion, the presence of virulence factor genes does not play a role in the histopathological degree of inflammation, the presence of bacteria was found to vary. Serum GPx activity increased in VF+ animals. While the hormone receptor expressions were similar, AQP expression was upregulated in the absence of virulence factor genes.
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    The role of nutritional-immunological indices in estimating serum LPS and antioxidant enzyme activity and sepsis status in female dogs with pyometra caused by E. coli
    (Elsevier, 2023) Yazlik, Murat Onur; Mutluer, Ipek; Kaya, Ufuk; Ozkan, Huseyin; Mustak, Inci Basak; Colakoglu, Hatice Esra; Altinbas, Yunus Furkan
    The aim of this study was to diagnose pyometra and related sepsis status using cost-effective nutritional-immunological indices, antioxidants, and toxin levels in dogs and to investigate the utility of the indices in predicting toxin and antioxidant status. A total of 29 dogs were enrolled into the present study. Among these, 9 female dogs in their diestrus stages, were allocated for elective ovariohysterectomy. The pyometra group was also separated into two subgroups as Sepsis (+) and Sepsis (-). Blood samples were collected into two tubes containing EDTA for hematological analysis; without anticoagulant for serum progesterone, LPS concentration, and antioxidant levels at the time of diagnosis. Bacteriological and tissue samples of the uterus were collected after the ovariohysterectomy. Antioxidant activity, progesterone, and toxin concentration were determined by using commercial ELISA kits. Statistical analyses were performed using Stata version 16.1 and MedCalc 16 statistical software. Receiver operating characteristics curves were used for the threshold for evaluating pyometra and sepsis status. Pairwise comparisons were carried out of the area under the curve (AUC) for thresholds of nutritional immunologic indices (hemoglobin, albumin, lymphocyte, platelet (HALP) score; prognostic nutritional index (PNI); Albumin hemoglobin index (AHI)), serum LPS and antioxidant activity. Linear regression model was used for the estimation of serum LPS and antioxidant activity by using indices. Mean serum progesterone, LPS concentrations, and Nitric Oxide (NO) production were greater, while serum superoxide dismutase (SOD), tissue SOD, and glutathione peroxidase (GPx) activities were lower in dogs with pyometra. All nutritional-immunologic indices were lower in pyometra cases. Nutritional-immunologic indices (AUC of HALP:0.759; PNI:0.981; AHI 0.994), NO (AUC: 0.787) and SOD (AUC: 0.784) levels were useful for pyometra diagnosis. AHI and LPS were useful for the determination of sepsis status with the AUC values of 0.850 and 0.740, respectively. While AHI was useful for the estimation of serum LPS and NO concentration (p < 0.001), PNI was useful for serum SOD concentration (p = 0.003). In conclusion, PNI, HALP and AHI can be used in the diagnosis of pyometra, however, only AHI and LPS levels can be used in the diagnosis of sepsis. SOD and NO can be used to determine pyometra but have no effect on determining sepsis status. Additionally, the estimation of the levels of serum LPS, NO, and SOD activities can be done using the AHI and PNI values.