Paršavedžių genitalinio trakto uždegimų etiologija, prevencija ir gydymas ; Inflammation etiology, prevention and treatment of genital tract in sows
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The research work has been carried out in the Lithuanian Veterinary Aca-LITHUANIAN VETERINARY ACADEMY demy, in 2002-2006, Kaunas, Lithuania. Research supervisor – Assoc. Prof. Dr. Eugenijus Aniulis (Biomedical Sciences, Veterinary Medi-cine – 12B) Lithuanian Veterinary Academy. Research advisers: Assoc. Prof. Dr. Albina Aniulien ė (Biomedical Sciences, Veterinary Medi-cine – 12B) Lithuanian Veterinary Academy; Dr. Raimundas Mockeli ūnas (Biomedical Sciences, Veterinary Medicine – 12B) Lithuanian Veterinary Academy. Chairman of Veterinary Medicine Science Council – Prof. Habil. Dr. Henrikas Žilinskas (Biomedical Sciences, Veterinary Medi-cine – 12B) Lithuanian Veterinary Academy. Žaneta Laureckien ė Members: Prof. Dr. Antanas Sederevi čius (Biomedical Sciences, Veterinary Medicine – 12B) Lithuanian Veterinary Academy; Prof. Habil. Dr. Vytautas Špakauskas (Biomedical Sciences, Veterinary Medicine – 12B) Veterinary Institute of Lithuanian Veterinary Academy; Prof. Dr. Bronius Bakutis (Biomedical Sciences, Veterinary Medicine – INFLAMMATION ETIOLOGY, 12B) Lithuanian Veterinary Academy; PREVENTION AND TREATMENT OF Dr. Violeta Juškien ė (Biomedical Sciences, Zootechny – 13B) Institute of GENITAL TRACT IN SOWS Animal Sciences of Lithuanian Veterinary Academy. Opponents: Prof. Habil. Dr. Algimantas Matusevi čius (Biomedical Sciences, Veterinary Medicine – 12B) Lithuanian Veterinary Academy; Prof. Habil. Dr.
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LITHUANIAN VETERINARY ACADEMY
aneta Laureckien INFLAMMATION ETIOLOGY, PREVENTION AND TREATMENT OF GENITAL TRACT IN SOWS Summary of Doctoral dissertation Biomedicine Sciences, Veterinary Medicine (12B) Kaunas, 2006
The research work has been carried out in the Lithuanian Veterinary Aca-demy, in 2002-2006, Kaunas, Lithuania. Research supervisor Assoc. Prof. Dr. Eugenijus Aniulis (Biomedical Sciences, Veterinary Medi-cine 12B) Lithuanian Veterinary Academy. Research advisers: Assoc. Prof. Dr. Albina Aniulien (Biomedical Sciences, Veterinary Medi-cine 12B) Lithuanian Veterinary Academy; Dr. Raimundas Mockeli nas (Biomedical Sciences, Veterinary Medicine 12B) Lithuanian Veterinary Academy. Chairman of Veterinary Medicine Science Council Prof. Habil. Dr. Henrikas ilinskas (Biomedical Sciences, Veterinary Medi-cine 12B) Lithuanian Veterinary Academy. Members: Prof. Dr. Antanas Sederevi č ius (Biomedical Sciences, Veterinary Medicine 12B) Lithuanian Veterinary Academy; Prof. Habil. Dr. Vytautas pakauskas (Biomedical Sciences, Veterinary Medicine 12B) Veterinary Institute of Lithuanian Veterinary Academy; Prof. Dr. Bronius Bakutis (Biomedical Sciences, Veterinary Medicine 12B) Lithuanian Veterinary Academy; Dr. Violeta Jukien (Biomedical Sciences, Zootechny 13B) Institute of Animal Sciences of Lithuanian Veterinary Academy. Opponents: Prof. Habil. Dr. Algimantas Matusevi č ius (Biomedical Sciences, Veterinary Medicine 12B) Lithuanian Veterinary Academy; Prof. Habil. Dr. Ramutis Klimas (Biomedical Sciences, Zootechny 13B) iauliai University. Public defence of Doctoral thesis in Veterinary Medicine Science Council will take place at Lithuanian Veterinary Academy I auditorium 14 am on 29 th November, 2006. Address: Til s 18 LT-47181, Kaunas, Lithuania. The abstract of doctoral dissertation has been send on 27 th of October, 2006 according to the confirmed address list. This dissertation is available at the libraries of the Lithuanian Veterinary Academy and LVA Veterinary Institute. 2
LIETUVOS VETERINARIJOS AKADEMIJA aneta Laureckien PARAVEDI GENITALINIO TRAKTO UDEGIM ETIOLOGIJA, PREVENCIJA IR GYDYMAS Daktaro disertacijos santrauka Biomedicinos mokslai, veterinarin medicina (12B) Kaunas, 2006
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Darbas atliktas 2002-2006 metais Lietuvos veterinarijos akademijoje. Mokslinis vadovas Doc. dr. Eugenijus Aniulis (Lietuvos veterinarijos akademija, biomedicinos mokslai, veterinarin medicina 12B). Mokslinio darbo konsultantai: Doc. dr. Albina Aniulien (Lietuvos veterinarijos akademija, biomedicinos mokslai, veterinarin medicina 12B); Dr. Raimundas Mockeli nas (Lietuvos veterinarijos akademija, biomedici-nos mokslai, veterinarin medicina 12B). Disertacija ginama Lietuvos veterinarijos akademijos Veterinarin s medici-nos mokslo krypties taryboje: Pirmininkas Prof. habil. dr. Henrikas ilinskas (Lietuvos veterinarijos akademija, bio-medicinos mokslai, veterinarin medicina 12B). Nariai: Prof. dr. Antanas Sederevi č ius (Lietuvos veterinarijos akademija, biomedi-cinos mokslai, veterinarin medicina 12B); Prof. habil. dr. Vytautas pakauskas (Lietuvos veterinarijos akademija, biomedicinos mokslai, veterinarin medicina 12B); Prof. dr. Bronius Bakutis (Lietuvos veterinarijos akademija, biomedicinos mokslai, veterinarin medicina 12B); Dr. Violeta Jukien (Lietuvos veterinarijos akademijos Gyvulininkyst s institutas, biomedicinos mokslai, zootechnika 13B). Oponentai: Prof. habil. dr. Algimantas Matusevi č ius (Lietuvos veterinarijos akademija, biomedicinos mokslai, veterinarin medicina 12B); Prof. habil. dr. Ramutis Klimas (iauli universitetas, biomedicinos mok-slai, zootechnika 13B). Disertacija bus ginama vieame Veterinarin s medicinos mokslo krypties tarybos pos dyje 2006 m. lapkri č io 29 d. 14 val. Lietuvos veterinarijos akademijos I auditorijoje. Adresas: Til s 18 g., LT-47181 Kaunas. Disertacijos santrauka isiuntin ta 2006 m. spalio 27d. Disertacij ą galima peri r ti Lietuvos veterinarijos akademijos ir LVA Vet-erinarijos instituto bibliotekose.
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INTRODUCTION Swine breeding - a branch of livestock breeding which supplies residents with food products and raw materials for industry. The success of swine reproduction is measured according to the number of piglets a sow produces per year, fertilisation index and size of litter. In order to achieve an optimal reproduction level, the sows anatomical and physiological systems must function well. A general understanding of basic swine anatomical and physiological reproduction systems can assist growers in foreseeing and eliminating possible disorders as well as easing decisions which affect swine breeding and related economic indicators. A large swine prolificacy, short gestation duration and rapid growth rate using minimal amounts of work expenditures allow for a large production amount. Lithuanian large hog complexes produce approximately 50% of pork. Having a large concentration of animals in hog complexes, sow prolifi-cacy is about 60-70%. The primary causes are due to sows affected by re-productional organ diseases, and post weaning low fertilization rate often leading towards culling. Statistical data shows that in hog complexes 10.6 47.7% are affected by reproductional organ inflammation. Post-partum inflammation processes are caused by microbes which enter the uterus via the vagina, urinary organs or digestive tract. The timely use of available veterinary prophylactic and treatment preparations in post-partum sows safeguards their healthiness and fertilization post-weaning. Tarasiul et al (1986) and Martineau (1992) suggest using parenteral anti-biotic applications for prophylaxis and treatment. There are literature re-sources (Dial and MacLacktan, 1988b; Bertschinger,1997, 1999a,b; Markowska-Daniel, 2001), which indicate that in treating post-partum re-productional organ inflammations, the use of systemic treatments is advis-able. Aims of the study: To determine the cause of post-partum reproductional tract inflammation and the influence of estradiol 17 β and progesterone during parturition. Also, to confirm available veterinary preparations 2.5% Sol. Cobactan®, 5% Sol Enroxil, Metricure® and Clamoxyl ® Metritis efficacy in prophylaxis and treatment of reproductional tract in-flammations. Goals of the study: 1. Determine the prevalence of genital tract inflammation in large hog complexes;
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2. Identify vaginal microflora in gilts and sows during estrus; 3. Ascertain sow vaginal microflora and its role in inflammation etiol-ogy 1-2 days prior to parturition; 4. Determine post-partum cervical cranial region microflora composi-tion and its role in inflammation etiology; 5. Compare the accuracy of chlamydial diagnotic methods; 6. Determine post-slaughter genital tract microflora in healthy purebred gilts and culled sows. Determine morphological changes of uterine mucosa in culled sows; 7. Determine the dynamics of estriodol 17 β and progesterone ranges in blood during parturition; 8. Determine the prophylactic efficacy of feed supplements Spectinomix 110, Lincosint 110 and veterinary preparation 5% Sol. Enroxil in treating reproductive tract inflammation; 9. Determine the prophylactic and treatment efficacy of 2.5% Sol. Co-bactan®, Metricure® ir Clamoxyl ® Metritis in post-partum uterine in-flammations. Novelty of the study: 1. The prevalence of sow reproductive organ disease was ascertained in the Lithuanias Republics large hog complexes; 2. The dominating microflora in sow reproductive organs and its influ-ence in post-weaning fertilisation were established; 3. The influence of prophylactic use of recommended feed supplements Spectinomix 110 and Lincosint 100 on reproductive organ microflora and piglet to weaning health was evaluated; 4. The prophylactic and treatment efficacy of veterinary preparations 2.5% sol. Cobactan®, Metricure®, Clamoxyl ® Metritis was established in sows with post-partum uterine inflammations. Practical assignment significance: A reproductive organ disease morbidity analysis was performed in large hog complexes. Attempts of feed supplements Spectinomix 110 and Lin-cosint 110 and their influence on sow reproductive tract microflora and pig-let health to weaning was performed. An evaluation was performed of vet-erinary preparations 5% Sol. Enroxil, 2.5% sol. Cobactan®, Metricure®, Clamoxyl ® Metri-tis and their influence on prevention and treatment sows reproductive tract inflammation. Volume and structure of the study: The dissertation was written in Lithuanian. It includes 102 pages; 12 headings, 9 tables, 23 illustrations, 136 literature references and an appendix (6 tables).
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RESEARCH METHODS Time, location and conditions of the research Investigations were performed during 2002-2006 at the Lithuanian Vet-erinary Academy, Dept. of Non-contagious Diseases, Obstetrics and Gyne-cology Division, Animal Reproduction Laboratory, System Hog Complex, Ltd. Data regarding sow genital tract disease morbidity was collected and summarized from 19 large hog complexes throughout Lithuania. Caudal vaginal smear samples were collected from all sows using single-use trans-portable media (Invasive sterile, EUROTUBO, Mfg. I.A.S.A., Spain). Cer-vical samples for bacteriological testing were collected 1 3 days before parturition and on 4 5 d of after treatment with 2.5% Cobactan, Metricure and Clamoxyl® Metritis using single-use sterile pipettes. Seeding was per-formed on McConkey (McConkey agar, Oxoid, England), Mannitol Salt Agar (E&O Laboratories Ltd, Scotland), Columbia Agar Chocolate (E&O Laboratories Ltd, Scotland), Strept. Sel.C.O.B.A. (E&O Laboratories Ltd, Scotland).The plates were incubated for 24-48 h at 37ºC under aerobic con-ditions. Every 24 h the grown colony size and their colour were evaluated. Grown colonies were tested with 3% hydrogen peroxide solution. To iden-tify Staph. aureus we used the latex kit, Staphytect Plus Test DR 850 (Oxoid, England). Bacteriological investigations were performed at the Lithuanian Veterinary Academy, Animal Reproduction Laboratory. Stock gilt and sow genital tract microflora testing during estrus For this study 23 stock gilts and 84 sows that were in estrus 5-7 days post-weaning were selected. Sows which were in estrus 5-7 days post-weaning were divided into 4 groups: 18 gilts, 15 2 nd breeding, 31 3 rd breed-ing and 20 multi-bred sows. Vaginal samples from 101 sows 1-2 days pre-parturition were bacte-riologically tested. Samples were obtained using single-use transportable media (Invasive sterile EUROTUBO Manufacturer I.A.S.A., Spain). Post-partum testing of cranial region of cervix microflora In seeking to identify post-partum uterine microflora composition, sam-ples were obtained for bacteriological testing from the cranial region of the cervix from 63 sows using apparatus PLM ( V.Gabrijolavi č ius, 1987). Examination of uterine mucosa morphological changes For this examination 3 sows which did not successfully fertilise post-partum were slaughtered. The uterine mucosal samples were fixed in 10% neutral formalin solution. The samples were prepared according to O.
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Kublickienes (1978) method. Estradiol 17 β and progesterone blood value variation dynamics dur-ing parturition For this experiment we selected 5 gilts, 5 bi para and 5 tri para sows. At post-parturition of the 1 st piglet, we obtained 10ml of blood via vacutainer from the sows ear vein. The 2 nd sample was obtained 1h after parturition of the 1 st piglet and the 3 rd sample was obtained after another hour. Blood se-rum was centrifuged for 15min. at 3000rpm. Micro test tubes were filled with 0.5 1ml blood serum and refrigerated to -20°C. A total of 45 samples were obtained. The Estradiol 17 β test was performed using the diagnostic kit E2-RIA-CT (BioSource Europe S.A., Belgium). Progesterone amounts were determined using the diagnostic kit PROG-RIA-CT (BioSource Europe S.A., Belgium.) Testing was performed at the Kaunas Medical University Clinic, Endo-crinology Institute, Radioimmune Testing Laboratory according to Duchens et al., (1994) methodology. Measurements were obtained via the Radioim-mune Analyzer ГАММА 12(Russia). Prophylaxis of sow reproductive organ inflammation For the investigation 90 sows were selected. They were divided into 3 groups of 30 each. In Group 1, 15 sows with 10 days remaining to parturi-tion received in feed (100kg 150gr) Spectinomix 110 (spectinomycin sul-phate 110g, filled to 1000g, Sintofarm, Italy), antibiotic additive and were fed this way until parturition. The Control Group did not receive feed addi-tives. In Group 2, 15 sows with 10 days remaining to parturition received feed antibiotic Lincosint 110 (linkomycin hidrochloride 110g, filled to 1000g, Sintofarm Italy). The Control Group did not receive feed additives. Group 3 sows received daily IM injections - 5 ml 5% Sol. Enroxil 3 days remaining until parturition. The preparation was not used in Control Group sows. Cranial region cervical samples for bacteriological testing were taken from the Experimental and Control Group sows on the day following partu-rition, using apparatus PLM (V.Gabrijolavi č ius, 1987). A total of 90 sam-ples were obtained.
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Chlamidial evaluation Using CF (Complement fixation) and ICF (Indirect complement fixa-tion) serologic reactions according to V.N. Siurin et al. (1986) methodology, 129 sows blood serum samples were tested for chlamidia. From the same 129 sows vaginal smears were obtained and direct immunofluorescence re-actions (IF) were performed. The smears were valued as positive which showed apple-green coloured interstices within the cells. Post slaughter stock gilt and sow urogenital organ microflora For microfloral testing from uterine horns, vaginal - near cervical ventral region and urinary bladder, we selected 13 healthy stock sows and 13 sows culled due to unsuccessful fertilization. Post-slaughter samples were ob-tained using single-use transportable media (Invasive sterile EUROTUBO Manufacturer I.A.S.A. Spain). Obtained samples were placed in an insulated carrier. The samples were sown on nutritive media 4-6 hours later. Use of veterinary preparations treating reproductive organ inflam-mations, prophylaxis and treatment efficacy Tests were conducted using 56 sows (42 Experimental and 14 Control), which showed signs of reproductive organ inflammation 1-2 days post-partum. Group 1 sows (14 Experimental) received 2 consecutive daily IM injec-tions of 2.5% Sol. Cobactan® (cefquinome sulfate, equivalent of 25 mg, cefquinome 26.64 mg, excip. to 1 ml; Intervet International GmbH, Ger-many). Group 2 received 4ml/100kg daily IM injections of 2.5% Sol.Cobactan® for 2 consecutive days and an intrauterine preparation Met-ricure® (cephapyrine 500mg, excip. to 19 g; Intervet International B.V., Holland). Group 3 received single-dose intrauterine applications of Clamoxyl ® Metritis (amoxicillin tryhydrate 0.84g, excip. to 21.30g (Pfizer Animal Health, Belgium). The Control Group sow reproductive organ invo-lution was limited to observation. Samples were taken from the cervix of Experimental Group sows for bacteriologic testing 4-5 days post treatment. The efficacy of used preparations was decided by changes within the uterine microflora, refertilisation, numbers of live and stillborn piglets, weight of newborn litter and weight of weaned piglets.
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RESEARCH RESULTS Incidence of sow genital tract inflammation In analysing the morbidity of sow reproductive organ diseases in 19 hog complexes in our Republic, we determined that during 2002 28,614 sows 13.7% - were registered with reproductive organ inflammations and during 2003 - 29,746 sows showing a morbidity rate of 10.6%. Vaginal microflora during estrus In examining vaginal mucosal samples from stock gilts during estrus, bacteriological pure cultures were grown 52.2%, ( Streptococcus spp.-75.0%, KNS 25.0%), mixed cultures 21.7% ( Streptococcus spp., KNS, Enterobacteriaceae ) and 26.1% samples were sterile.
38,90% 39,10% 36,60% 25,00%
0,00% 10,00% 20,00% 30,00% 40,00% 50,00% Para I Para II Para III Para IV Fig. 1. S. aureus identified from vaginal secretions of various-aged sows 1 pav. vairaus amiaus rujojan č i paravedi makties sekrete iskir-tas S. aureus Para I gilt samples grew pure microbe cultures - 33.3% ( Streptococcus spp.), mixed - 66.7%. ( Streptococcus spp . , KNS, Enterobacteriaceae spp. ) 10
25% of the mixed microflora composition S. aureus was identified. Para II sows pure microbe cultures were obtained from 26.7% of the samples ( Streptococcus spp. ) and mixed cultures 73.3% ( Streptococcus spp., KNS, Enterobacteriaceae spp. ) 73.3 %. In 36.6% of the mixed cultures S. aureus was identified . Para III sows, vaginal secretion pure cultures were dominated by Streptococcus spp. (25.8 %), and mixed microflora ( Strepto-coccus spp., KNS, Enterobacteriaceae eima) was identified 74.2%. In the mixed microflora composition S. aureus was identified 39.1%. Para IV sow vaginal secretion pure cultures were grown in only 10.0% -( Streptococcus spp .), mixed - 90% ( Streptococcus spp., KNS, Enterobacte-riaceae spp. ). In mixed cultures S. aureus (38,9%) was identified. Vaginal microflora in sows 1-2 days prior to parturition. With 1-2 days remaining to parturition, vaginal secretion grew pure cul-tures in 23.76%, of cases, comprised of ( Streptococcus spp. 16.67% and Enterobacteriaceae spp 83.33%). Mixed microbe cultures were identified in 61.4% of samples, comprised of Streptococcus spp., KNS, Enterobacteri-aceaespp. No microbe cultures grew in 14.85%.
14,85%
23,76%
61,39%
Mixed culture Samples culture Notably Fig.2. Vaginal secretion microflora 1-2 prior to parturition 2.pav. Makties sekreto mikroflora likus 1-2 paroms iki apsipariavimo Microflora from the urogenital tract of postmortem healthy and re-productive organ inflammation affected sows. Postmortem examination of stock gilts and culled sows urogenital tract microbial contamination showed that stock gilt uterine secretions were ster-ile, however samples taken from the urinary bladder grew pure cultures in 40% of cases ( Streptococcus spp. ), mixed 60% ( Streptococcus spp. , KNS, 11
Enterobacteriaceae spp. ). Samples from the vaginal cranial region grew mixed microflora in 100% of cases ( Streptococcus spp., KNS, Enterobacte-riaceae spp. ). Mixed cultures of Streptococcus spp., KNS, Enterobacteriaceae spp., of which 38.5% S.aureus was identified grew in 100% of vaginal secretion samples obtained from sows culled due to unsuccessful refertilisation. Sow urinary bladder secretion grew 30.8% pure Streptococcus spp. cul-tures and 69.2% grew mixed cultures ( Streptococcus spp., Enterobacteri-aceae family, KNS). All samples of sow vaginal secretion samples obtained from the cranial region grew mixed cultures of which 53.8% S.aureus. was identified. 100,00% 100,00% 90,00% 80,00% 70,00% 60,00% 60,00% 50,00% 40,00% 40,00% 30,00% 20,00% 10,00% 0,00% Uretra Vagina Uterus Mixed culture Sam ples culture Fig.3. Stock gilt urogenital tract microflora post slaughter 3.pav. Veislini kiaulai č i urogenitalinio trakto mikroflora po pasker-dimo Morphological changes of diseased sow uterine mucosa The uterine content in slaughtered sows was dirty and grayish yellow. The endometrium was congested and swollen, small laminal haemorrhages and eosinophilic infiltration in all parts of the endometrium including glands and blood vessels was observed. Uterine glands were depleted and dilated. 12
Post-slaughter examinations of sow reproductive organs indicated exu-date and macro changes of endometrium. The diagnosis (chronic endometri-tis) was confirmed via histological examination of uterine mucosa. Periglandular fibrosis and leukocytosis was noted, with predominant lymphocytes. The endometrium was congested, lumen vessels included lymphocytes and neutrophils. Accumulations of neutrophils and eosinophils were found in the stroma and glands. Granules of haemosiderin were in the stratus compactum . The glands were depleted and those which survived were atrophic, flattened, and reduced or cystic. Dilation of lymphatics in the stratum compactum and lumina epithelium were evident. Estradiol 17 β and progesterone blood value variation dynamics dur-ing parturition Our examinations show that estradiol 17 β in Para I gilts was 10.279±0.53pmol/l which is 2.603pmol/l less than in Para II sows and even 3.082pmol/l less than in Para III sows. Two hours post-parturition the lowest estradiol 17 β amount was in 1 st breeding gilts (7.970±1.15pmol/l), 1.121pmol/l in 2 nd breeding and 1.261pmol/l in 3 rd breeding sows. During parturition, the progesterone amounts gradually decreased in all sows. Estradiol 17 β and progesterone blood values were induced by the du-ration of parturition and number of stillborn piglets. Para I gilts had the low-est blood values of estradiolo 17 β during parturition, and had conditionally more stillbirths than Para II and Para III sows. Sow cervical secretion microflora 1-2 days post-partum In examining sow cervical secretions 1-2 days post-partum, 20.63% samples were sterile, 23.82% - grew pure cultures and 55.55% - mixed mi-crobe cultures. Pure microbe cultures were comprised of Streptococcus spp.-53.84% and Enterobacteriaceae spp. 46.16%. Mixed microbe cultures were comprised of Enterobacteriaceae spp. 22.86%, Streptococcus spp.- 40.00% and KNS 37.14%. Chlamidia identification In examining sows using three comparative methods (CF, ICF, IF) we determined that using serological CF, chlamidia was diagnosed in 11.6%, ICF 16.3% and IF 22.5% of obtained samples. Comparison of post-partum prophylactic effect of feed additives Spectinomix 110, Lincosint 110 and preparation 5% Sol. Enroxil on reproductive organ inflammation
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With 10 days remaining to parturition, experimental group sows received feed additives of Spectinomix 110 and Lincosint 110, and their vaginal se-cretions obtained 1 day post-partum grew pure and mixed microbe cultures. The same species ( Enterobacteriaceae spp. , Streptococcus spp. , KNS) were identified in both experimental and control group sows. In using Spectinomix 110, post-partum cultures grew pure in 52.0% of cases and 48.0% - mixed. Respectively, the Control Group 56.0% pure and 44.0% mixed. Using Lincosint 110 in the Experimental Group, pure microbe cultures were identified in 53.3% of cases and mixed 46.7%. The Control Group sow samples grew 46.7% pure, and 53.3% mixed cultures. Table 1. Prophylactic efficacy of preparations Spectinomix 110, Lin-cosint 110 and 5% Sol. Enroxil Live pig- Stillborn Weaned Num- Total lets piglets piglets Sows er of Preparation pig-sows lets Nr. % Nr. % sk % Ex-pmeerni--15 Spec1ti1n0omix 176 16292 14 8 157 96.9 I tal tCrooln -15 - 180 15684.6 24 15.4131 83.9 Ex-pmeerni--15 Lin1c1o0sint 159 15194.968 5.04143 94.7 II t l a tColn-15 - 160 14892.5 12 7.5 132 89.2 ro Ex-III tpmelerni--15 5% rSooxli.l En-170 16597 5 3 157 95.15 a Con- 15 - 164 149 90.85 15 9.15 132 80.49 trol I p ≤ 0.037; II p ≤ 0.082; III p ≤ 0.001
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Thre days prior to parturition the sows received daily IM injections of 5ml 5% sol. Enroxil. One day post-partum samples of vaginal secretions were sterile. In comparison the Control Group vaginal secretion samples grew 53.3% pure, and 46.7% mixed microbe cultures. Tests show that antibiotic feed additives Spectinomix 110 and Lincosint 110 did not have an influence on vaginal microflora post-partum, however there were more healthy weaned piglets than in the Control Group (respec-tively p<0.037 and p<0.082) During the 3 rd investigation, prophylactically injecting 5% Sol. Enroxil to Experimental Group post-partum sows not only destroyed vaginal micro-flora but also increased the number of healthy weaned piglets by 14.66% (p ≤ 0.001) over the Control Group. Efficacy of veterinary preparations 2.5% Sol. Cobactan ®, Met-ricure® and Clamoxyl ® Metritis in sow uterine inflammation prophy-laxis. Bacteriological examinations of vaginal smears taken before parturition showed mixed microflora in 85.4%, including: Enterobacter sp .-42.9% , Streptococcus sp. 57.1%, Staphylococcus sp. 62.3%. Pure cultures ( Staphylococcus sp) were evidenced in 14.3% sows. In 50% uterine sam-ples, taken 4 5 d after treatment from group I sows, grew Enterobacter sp. in pure cultures. Mixed microflora ( Enterobacter sp . and Streptococcus sp.) were found in 50%. In cervical smear samples from group II - 42.9% pure cultures ( Enterobacter sp . -14.3% and Staphylococcus sp. 28.6%) were isolated. From samples in this group mixed cultures grew only in 14.2% ( Enterobacter sp . -7.1% and Staphylococcus sp . - 7.1%). No cultures grew in 42.9% of the samples. In cervical smear samples from group III sows no growth was observed in 41.4%. Pure cultures were identified in 37.1% ( Enterobacter sp . 14.3%, Streptococcus sp. 22.8%) and mixed micro-flora in 21.4% ( Enterobacter sp .-7.1% , Streptococcus sp. 7.1%, Staphylo-coccus sp. 7.2%.). In the control group (IV) from the cervical smears after farrowing mixed microflora in 85.7%, and pure culture in 14.3% of the samples were found. Prevention of reproductive organ inflammation had an influence on the health of the piglets. The average weight of the litter was larger in the test groups in comparison to the control group (Table 1). This was particularly evident in Group II, where Cobactan was com-bined with Metricure. Successful post-weaning fertilization required a dif-ferent number of insemination doses. The Control Group required 1.64 ± 0.23 semen doses, where as in Cobactan (Group I) 1.5 ±0.52 doses, Co-bactan and Metricure (Group II) 1.0 dose, and intrauterine Clamoxyl® Metritis (group III) also 1 AI. Comparing the used preparations with the control group, we noticed that the fertilization data obtained from the test 15
groups II and III were statistically significant ( 0.034 and 0.024). Table 2. Comparison of efficiency of preparations Prepara- Num- Route AI Num- Num- Mean Num- Mean tion er of of ad- index ber of ber of weigh ber of weigh sows minis- piglets piglets t of piglets t of a tration born born deliv- at litter alive dead ered wean- at (kg) ing wean-ing .5 % 61.79 9.93 I t2Caonb ® ac14 i. m. ±01..55 2±100..9856 ±01..9539 ±131..6545 ±6.68 ±0.92 - 2.5 % Cobac-m. 10.57 0.2 0.57 II taannd ® 14 i.i.u. 1.0 ±0.51 ±0.691 ±103..7876 7±57..0600 1±0.51 Met-ricure ® III Cll ® amox14 i.u. 1.0 ±100..7997 ±00.71 14.21 69.64 10.43 yMe tritis .99 ±1.19±5.71 ±1.02 IV Control 14 1.64 10.50 0.93 12.93 61.29 9.57 - ±0.84 ±1.51 ±1.14 ±1.69 ±8.93 ±1.28 RESULT DISSCUSSION Swine-breeding is one of the most developing areas of livestock breed-ing in Lithuania. Lately, approximately 50% of all pork produced comes from large hog complexes. As the concentration of animals increases, it be-comes more challenging to provide optimal husbandry, feeding and care conditions. In large swine-breeding complexes, the role of timely prophylaxis and treatment preventing post-partum reproductional organ inflammation be-comes increasingly important. In analyzing the morbidity of sow post-partum uterine inflammation in 19 swine-breeding complexes, it was determined that during 2002, an aver-age of 2.974 (10.6%) sows were affected out of 29.749. Though that is not a high morbidity rate, in other complexes it was 20.4 % - 47.7%. Other au-thors also write about a rather high uterine inflammation morbidity rate (Bilkei et al.1995; Waller et al. 2002). Their published statistical data is 16
similar to our analysis results. Microflora is constantly found in the vaginal vestibule, vagina and distal region of the urinary bladder of healthy stock gilts and sows. The urinary bladder is usually sterile. In examining urine, microbes are found which arise from the urethra or vaginal vestibule (Moller et al., 1981; Berner, 1988; Maes et al.1999). The research of Bave et al. (1993) shows that the variation of cervical and vaginal microbes depends on the individual sows reproductional tract mechanism hormonal cycle, immunoglobulin and mucosal secretions, granulocyte phagocytic activity. McLean et al. (1974) indicates that parturi-tion and changes in feed ration influence E.coli and other bacteria reproduc-tion in the porcine urogenital tract. His data shows that vaginal microbial contamination in older sows is larger. In our research we tested microbial contamination of stock gilt and sow mucosal secretions during estrus. Bacteriological testing of 23 stock gilt mucosal samples grew 52.2% pure cultures, 21.7% mixed and 26.10% grew no cultures. Pure and mixed cultures were comprised of Streptococcus spp. , Enterobacteriaceae spp. and KNS. A large contamination was noted in Para I gilts. 33.3% samples grew pure cultures and 66.7% - mixed. In 25% of the mixed microflora cultures S.aureus was identified. As parturition numbers increased (Para II) so did vaginal mucosal bacte-rial contamination. In this group of sows, in comparison to Para I sows we identified 6.6% less pure, though 6.6% more mixed cultures. Notably, in mixed microbe composition 33.6% samples contained S.aureus . Investigation the vaginal mucous samples of Para III sows in estrus, 25.8% grew pure cultures and 74.2% grew mixed. Pure and mixed microbe cultures were comprised of Streptococcus spp. , KNS and Enterobacteri-aceae spp. microbes. Mixed microflora cultures of vaginal secretion samples identified S.aureus in 39.1% of cases. This contagious microbe was identi-fied 14.1% more often than in Para I gilts and 2.74% more so than in Para II sows vaginal secretions. Vaginal mucous samples from Para IV sows in estrus grew pure cultures in 10.0% of cases and 90.0% mixed. All pure and mixed cultures grown from vaginal mucous samples from sows in estrus samples contained Enterobacteriaceae spp. , Streptococcus spp. and KNS, however in mixed cultures from older sows S. aureus was identified. An increasing vaginal contamination of S. aureus was also de-termined by other authors (Maes et al. 1999; de Winter et al.2002). We observed that bacteriolocially testing vaginal secretions with 1-2 days prior to parturition, the same microflora is found as in during estrus.
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Pure microbe cultures comprised 23.76% and mixed 61.39%. In this ex-perimental group 14.85% grew no cultures. S.aureus was not identified ei-ther, coinciding with Bostedt et al., (1998); Zhao Jing et al. (1998) and Bertschringer (1999) data. However, there is data in literature stating that there is other microflora in pregnant and non-pregnant sow vaginas. Sanders and Bilkei (2004) indi-cate that stress prior and during parturition can predispose chronic urogeni-tal infections. Waller et al. (2002) noted that in large swine-breeding com-plexes with poor husbandry (microlimate, feeding, high density concentra-tion), sows were often affected by MMA. The passage of identified micro-flora into the vaginal vestibule, vagina and cervix is stipulated due to the sows sitting position (Berner, 1984). Sow uterine microflora is comprised of broad spectrum anaerobe and aerobe species. Bilkei et al., 1994 and Waller et al., 2002 indicate that most commonly Streptococcus spp., Staphylococcus spp., Corynebacterium spp., E.coli, Klepsiella spp. and Actinobacillus spp. are found. Most of these bac-teria cause urogenital tract inflammations showing as vaginal secretions (De Winter et al.1995). Maes et al. (1999) data indicates that the existance of microorganisms in the uterus does not always cause endometritis. Jubb et al.(1993) research establishes that non-pregnant sows uteri are resistant to infection. McEntee (1990) findings indicate that the cervix is an effective protection barrier against microbe entry. During estrus and parturition the cervix is open. Notwithstanding from the cervical barrier function, microbes can pass from the vagina into the uterus. Our research indicates that secretions tested from the cranial region of the cervix of 63 sows 1-2 days post-parturition, pure cultures were com-prised of Enterobacteriaceae spp. 46.16% and Streptococcus spp.-53.84%. Mixed microbe cultures were comprised of Streptococcus spp. -40.0%, Enterobacteriaceae spp. - 22.86% and KNS 37.14%. The species we identified were confirmed by De Winter (1995), Bilkei et al. (1995) and Waller et al. (2002) research. Their research indicates that the presence of microbes in the uterus not always causes endometritis nor early pregnancy partial embryo mortality. Authors in literature and our research confirms that microbes identified in post-parturition uteri can be potential causative inflammation agents due to disorders of uterine muscle rigidity. Another urogenital tract inflammation causative agent can be chlamidia (Chlamydie suis). They are a gram-negative reproducing cells. In primary foci, the disease manifests in pregnant sows and newborn piglets. Gerulis (2002) data indicates that clinical chlamidiosis appears under the influence
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of stress when husbandry and feeding practices are poor. We used 129 sows for our investigation and tested them via 3 methods (KSR, NKSR and TIF). We established variations in the testing method ac-curacy. Testing sow blood serum samples using the KSR method, a positive reaction was shown in 11.6% cases, NKSR method - 16.3% . In testing the same 129 sows using the TIF method, a positive reaction was achieved in 22.5% of cases. The test positive reaction confirms that chlamidia reproduce in the cell. Our data coincides with Bagdonas (1998), Sachse (2002) and Gerulis (2002) research. Examining microbiological and histopathological test samples from slaughtered sow reproductive tracts not only enables objective determination of lesions but aids in obtaining samples too. Miurhead (1986) indicates the necessity of urogenital tract microbiological and histopathological testing. Due to anatomical structure, obtaining uterine mucosa biopsy samples from live sows is complicated. The aforementioned author, having slaughtered 47 sows and in detail examined the urogenital tract, did not find any pathologi-cal uterine changes in notable 44% of cases. Seeking to examine stock gilt and sow non-fertility causes, we performed post-slaughter microbiological tests of the urogenital tract. Our data establishes that uterine secretion sam-ples obtained from 5 slaughtered stock gilts were sterile. However in sam-ples obtained from urinary bladders grew pure cultures ( Streptococcus spp. ) in 2 samples (40.0%) and mixed microbe cultures ( Enterobacteriaceae spp. , Streptococcus spp. and KNS) were identified in 3 samples (60.0%). Examin-ing microbiologically secretion from the cranial region of the vagina, mixed microflora was identified in all samples. ( Enterobacteriaceae family , KNS ir Streptococcus spp ). Examining 13 uterine secretion samples from non-fertilised and thusly culled sows, all samples grew mixed microflora. A notable 38.5% of mixed microbe cultures contained S.aureus. Examined urinary bladder secretios grew pure (30.8%) and mixed mi-crobe cultures (69,2%) Streptococcus spp. were identified in pure cultures and in mixed -Enterobacteriaceae spp. , Streptococcus spp. and KNS. S. aureus (53,8%) was identified in mixed microflora cultures from sam-ples obtained from the cranial region of sow vaginas. Our test results con-firm the results of earlier investigations. Dial et al. (1988); Dee, (1992); Wendt, (1998); Makovska-Daniel, (2001) and Stczubial et al. (2003) indi-cate that E.coli, S.aureus, Streptococcus spp., Pasteurella multocida, Klep-siella spp. and , Enterobacteriaceae spp. are usually identified in cultures grown from sow urogenital tracts. Our performed post-slaughter urogenital tract secretion microbacterial tests indicate that stock gilt uteri are sterile. However, samples obtained
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from the cranial region of the vagina or urinary bladder grow pure and mixed cultures. Uterine, urinary bladder and vaginal secretions obtained from culled sows grew saprophytic microflora alongside contagious S.aureus , which produces toxins, causes endometritis and thereby impedes normal fertilisation. Prophylactic IM antibiotic injections during late pregnancy can prevent post-partum reproduction organ inflammation. Based on literature data, prostaglandins are usually indicated for this purpose (Oesrtophan, Dynolitic and others.) According to K.Katowskis (2003) data, 2 weeks prior to expected partu-rition, it is necessary to prophylactically administer Evetsel injections (so-dium selenite and Vit. vit. E). The aforementioned authors data indicates that following Evetsel injections, sows do not show symptoms of mammary gland disease or vaginal secretions. The numbers of live births increase, litter weight is higher as is the number of weaned piglets. Our study data indicates that 10 days prior to parturition, the use of anti-biotic feed additives Spectinomix 110 and Lincosint 110 did not decrease the incidence of reproductive organ bacterial contamination, however the numbers of numbers of weaned piglets increased by 13% (p ≤ 0.037) and by 5.5% (p ≤ 0.082) in our Control Group. IM injections of 5% Sol. Enroxil 3 days prior to parturition not only decreased reproductive organ bacterial contamination but also increased the number of weaned piglets by 14.66% (p ≤ 0.01). Our study results coincide with Feldman et al.(1998) and Horstein et al. (1998) research. Parturition prolonged by more than 3-4 hours usually results in more stillborn piglets and uterine lochia discharge is delayed. When uterine tonic-ity is compromised, the existing microbes reproduce and produce endotox-ins which lead to inflammation. For treatment, antibiotics and/or other com-binations of antimicrobial preparations are usually administered. Szcubial et al.(2003) identified causative uterine inflammation agents and in performing an antibiogram established that treating against E.coli , S.aureus , KNS and Streptococcus spp. penicillin group preparations, Streptomycin, Linkomy-cin, Tetracycline and Amoxicillin combined with clavulanic acid is indi-cated. In treatment it is important to use antibiotics to which identified mi-crobes are sensitive. Treating sow inflammation, Markovska-Daniel (2001) used: Group 1 diseased sows received two daily 10ml intra-uterine applica-tions of Amoxiclav® and Group 2 sows received systemic treatment IM injections of Oxyytetracycline or Amoxicillin, and intra-uterine applications of Neomicine suspension. Her achieved research results confirm that Amoxiclav® is effective in treating MMA syndrome. Our performed micro-
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biological tests (42 experimental and 14 control sows) of obtained pre-partum vaginal secretions grew 86.0% mixed microbe cultures; Streptococ-cus spp. - 57.1%, Enterobacteriaceae spp. 42.9% and KNS 62.3%. Pure cultures of KNS were identified in only 14.0% of samples. Our research data is similar to that of Ross et al. (1983); Busse et al. (2002) and Szcubial et al. (2003). In treating sows affected by reproductive organ inflammations we com-pared 3 veterinary preparations: 2.5% Sol. Cobactan®, Metricure® and Clamoxyl ® Metritis, all having a strong antimicrobial effect. After adminis-tering 2.5% Sol. Cobactan® injections and obtaining cervical secretions 4-5 days post-partum, 50.0% samples grew pure ( Enterobacteriaceae spp. ) and 50.0% grew mixed microbe cultures comprised of Streptococcus spp. , En-terobacteriaceae spp. and KNS. Using systemic treatment (2.5% Sol. Cobactan® IM and Metricure® in-tra-uterine) and obtaining cranial region cervical secretion samples 4-5 days post-partum, pure cultures were identified in 42.9% of the samples com-prised of Enterobacteriaceae spp. and Streptococcus spp .. Mixed cultures grew in 14.2% of samples and notably 42.9% of the uterine secretion sam-ples were sterile. Sows treated with intra-uterine applications of Clamoxyl ® Metritis and samples obtained from the cervix grew pure cultures 37.2%, mixed 21.4%. A notable 41.4% proved sterile. In 4-5 days post-partum, cervical secretion samples were obtained from Control Group sows that grew pure cultures 14.3% , and mixed 85.7%. We can compare our study results with Gordzinski et al. (2001) treat-ment results. The aforementioned author treated sows affected with MMA syndrome via injections of Metrisan AN, which is a combination of Am-picillin, Neomycin and agents that increase phagocyte activity. During the course of treatment the author observed a quick lochia discharge and later a positive effect on fertilization. In using various antibiotics, a rapid healing was noticed by Wawron, (1997); Kryzanowski et al.(1998); Kotowski et al. (1998) and Gardzinski et al. (2001). However, the application of intra-uterine medicines is not always applied in treating cervical closure. This treatment method can be applied only post-partum or during estrus. Intra-uterine preparation applications is especially effective in the case of acute inflammation (Busse et al., 2002). The use of parenteral or IM antimicrobial preparations decreases uterine secretion microbial contamination, though it is quite important to note sow fertilization. That is shown via the insemination index (the number of in-seminations necessary for fertilisation). Sows that received prophylactic IM injections of 2.5% Sol. Cobactan® 21
had a lower insemination index (1.5±0.52) which was 0.14 less than the sows in the Control Group. The lowest insemination indexes were in those sow groups that had re-ceived prophylactic systemic metaphylaxis (2.5% Sol. Cobactan® IM and Metricure® intra-uterine) and in the other group single-dose intrauterine Clamoxyl ®Metrite. The insemination index of the sows in the Experimen-tal Groups was 1.0. That shows that sows in estrus after weaning their pig-lets were fertilized after the first insemination. In comparing the 2nd (2.5% Sol.Cobactan® and Metricure®), as well as the 3rd (Clamoxyl ®Metritis) sow group, the insemination index with the Control Group that is statisti-cally credible (<0.034 and <0.024). A better uterine lochal post-partum discharge is shown not only by the high insemination index, but alson by a lower number of stillborn piglets, a larger newborn litter weight, a larger weaned piglet number and their com-bined weight. CONCLUSIONS 1. Sows raised in large swine-breeding farms often suffer from post-partum reproductive tract disease (0.7% 87.2%). 2. Within stock gilt and sow vaginas, uteri and urinary bladders, Strepto-coccus spp. , Enterobacteriaceae spp. and KNS dominate in pure and mixed culture associations; 21.6% of vaginal mucosa samples obtained from gilts in estrus are notably sterile. 3. Vaginal mucosa samples obtained from Para I and older sows on es- trus grew mixed cultures including S.aureus (25.0% 39.1%). 4. The uterus of post-slaughter gilts is sterile in 26.10% of cases. How-ever, the urinary bladder and cranial region of the vagina grew pure (52.20%) and mixed microbe associated cultures (21.70%) which were comprised of Streptococcus spp. , Enterobacteriaceae spp. and KNS. S.aureus was identified (25%-39.10%) in reproductive organ samples that were obtained from Para I and older sows culled due to non-fertilisation. 5. Sows culled due to non-fertilisation showed morphological changes in the uterine mucosa: the subepithelial layers showed a strong lymphocyte and neutrophil infiltration, a flourishment of connective tissue was observed surrounding the glands which were deformed or atrophied. In the case of purulent endometritis, uterine purulence encompassing blood vessels and glands showed a high leucocyte infiltration. 6. Estradiol 17 β and progesterone blood ratios determined the duration of parturition and numbers of piglets born live. In Para I sows, lower blood values of estradiol 17 β amounts increased the number of stillborn by 1.2 1.4 in comparison to older sows. 22
