Isolation, identification and antibiotic resistance of Campylobacter strains isolated from domestic and free-living pigeons.

1. The aim of this study was to evaluate the occurrence of Campylobacter spp. in domestic and free-living pigeons and to evaluate the antibiotic resistance profiles. 2. The material consisted of cloacal swabs obtained from 108 homing pigeons and fresh faeces from 72 wild birds from Lublin and its vicinity. The identification of strains isolated on differential/selective media for Campylobacter spp. was carried out by MALDI-TOF and PCR. The susceptibility to antibiotics was evaluated by minimum inhibitory concentration (MIC) in Mueller-Hinton broth. 3. A total of 35 strains of Campylobacter spp. were isolated; 27 were identified as Campylobacter jejuni and 8 as Campylobacter coli. Over half of the isolates were resistant to erythromycin and streptomycin, 40% of strains were resistant to tetracycline and ampicillin and 37% isolates were resistant to amoxicillin. Resistance to two or more antibiotics was observed in all strains tested. 4. The results indicate that both domestic and free-living pigeons are reservoirs for bacteria of the genus Campylobacter, which are characterised by varied and growing resistance to commonly used antibiotics.

Antibiotic susceptibility of Lactobacillus strains isolated from domestic geese.

The aim of this study was to determine the antibiotic susceptibility of 93 Lactobacillus strains isolated from domestic geese raised on Polish farms. The minimal inhibitory concentration (MIC) of 13 antimicrobial substances was determined by the broth microdilution method. All strains were sensitive to the cell wall inhibitors ampicillin and amoxicillin (MIC ≤ 8 µg/ml). Resistance to inhibitors of protein synthesis and to fluoroquinolone inhibitors of replication was found in 44.1% and 60.2% of isolates, respectively; 26.9% strains were resistant to neomycin (MIC ≥ 64 µg/ml), 23.6% to tetracycline (MIC ≥ 32 µg/ml), 15% to lincomycin (MIC ≥ 64 µg/ml), 18.3% to doxycycline (MIC ≥ 32 µg/ml), 9.7% to tylosin (MIC ≥ 32 µg/ml), 56% to flumequine (MIC ≥ 256 µg/ml) and 22.6% to enrofloxacin (MIC ≥ 64 µg/ml). Bimodal distribution of MICs indicative of acquired resistance and unimodal distribution of the high MIC values indicative of intrinsic resistance were correlated with Lactobacillus species. Eleven (11.8%) strains displayed multiple resistance for at least three classes of antibiotics. Data derived from this study can be used as a basis for reviewing current microbiological breakpoints for categorisation of susceptible and resistant strains of Lactobacillus genus and help to assess the hazards associated with the occurrence of drug resistance among natural intestinal microflora.

Detection of the bovine viral diarrhoea virus (BVDV) in young beef cattle in eastern and south-eastern regions of Poland.

In view of the scarcity of information concerning viral diarrhoea virus (BVDV) infections in beef cattle in Poland, the aim of this study was to evaluate the presence of the BVDV in young beef cattle from selected herds in eastern and south-eastern regions of Poland. The material consisted of 78 sera obtained from beef cattle from 15 farms, aged 6-12 months. The anti-BVDV antibody level in the sera was estimated with an ELISA kit, and detection of the BVDV was carried out by standard PCR and one step Real-Time RT-PCR. The ELISA results showed a high degree (80%) of positivity in 5 of the 78 samples. In 7 samples the degree of positivity was in the very low range: < 40%. Of the 78 cDNA samples, the presence of genetic material with a length of 288 bp was found by standard PCR in 3 sera. The genetic material of BVDV was also found in the sera of the same three calves by Real-Time HRM PCR. BVDV infection in young beef cattle in south-eastern Poland is not a significant problem. This was confirmed by the positive ELISA results for 6.4% of the animals and the positive PCR results for 3.9%. The percentage of positive beef herds was about 8.6%. However, due to the severe nature of the disease and rapid transmission of the virus, regular monitoring of BVDV should be carried out.

An electrophoretic characterization of iron-transporting proteins in Mannheimia haemolytica A1.

Iron-regulated outer membrane proteins (IROMPs) in Mannheimia haemolytica A1, which function as a receptor for complexes containing iron ions, are induced by iron deficiency in the growth environment of the bacteria. Densitometric analysis of SDS-PAGE separation showed expression of IROMPs of 71, 77, and 100 kDa in the case of bacteria grown in a medium with 2,2-dipyridyl. The electrophoregrams obtained in 2-DE separations confirmed the presence of protein fractions with these molecular weights and isoelectric points ranging from 5.4 to 6.4. The results of the study also confirmed the ability of M. haemolytica A1 proteins involved in iron uptake to induce a protective immune response. In Western blot with serum from convalescent calves naturally infected with M. haemolytica A1, distinct reactions were obtained for IROMPs of 71, 77, and 100 kDa.

Effect of conglutinin on phagocytic activity of bovine granulocytes.

In the present study we investigated the effect of bovine conglutinin on the phagocytic activity of leukocytes. We measured both the chemotactic activity of conglutinin and its effect on the internalization of zymosan particles and E. coli by granulocytes. We also assessed the binding of conglutinin to various microorganisms isolated from clinical cases in cattle. We showed that conglutinin binds strongly to the surface of yeast cells and to mannan-rich zymosan particles, while weak binding was observed in the case of the bacterial strains tested, including those whose O antigen is composed of mannan. Conglutinin (1-10 microg/ml) neither acts as a chemotactic factor for peripheral blood leukocytes nor affects ingestion of E. coli by granulocytes. However, as flow cytometry based assay showed, conglutinin (0.1-1 microg/ml) increased ingestion of zymosan expressed as mean fluorescence intensity (MFI) of positive cells.

Analysis of the culture-derived soluble Babesia canis canis antigens derived from the Polish strains of the parasites.

UNLABELLED: OBJECTIVE, MATERIAL AND METHODS: The aim of this study was to analyse the protein fractions of the soluble parasitic antigen (SPA) from in vitro cultures of the native Polish strains of Babesia canis canis and to determine their immunogenicity through Western blotting using the sera of dogs vaccinated with this antigen. RESULTS: Polyacrylamide gel electrophoresis revealed 21 protein fractions with molecular weights from 12 to 205 kDa. The most intense reaction in Western blotting was observed between the serum antibodies of the SPA-vaccinated dogs and the fraction with the molecular weight of 52 kDa. CONCLUSION: Detailed studies on the composition of SPA of Babesia canis canis and reactivity of its individual protein fractions can be a starting point for the development of subunit vaccines against babesiosis. Using a preparation with only some electrophoretic fractions of SPA in the production of vaccines would allow to avoid putting an unnecessary protein burden in the vaccine which could cause side effects.

Characterization of outer membrane proteins participating in iron transport in Pasteurella multocida serotype A3.

Iron-regulated outer membrane proteins (IROMPs) of P. multocida serotype A3, which function as receptors for complexes containing iron ions, are induced by iron deficiency in the bacterial growth environment. Analysis of an electrophoresis image of proteins isolated from bacteria grown on medium supplemented with 2,2'-dipyridyl revealed expression of 16 new proteins that were not noted in the case of the bacteria grown in standard conditions, with molecular weights from 30 to 160 kDa. Induction of IROMP expression occurred within 30 minutes after restricted iron conditions were established. In immunoblotting, distinct reactions were noted for proteins of molecular weight ranges of 25-49 kDa, 61-95 kDa, and 108-214 kDa. Proteins of the molecular weight of 68, 75 and 86 kDa were analysed using mass spectrometry and matched with the highest probability to proteins in the NCBI data base. Several dozen different proteins with similar amino acid sequences were matched to each sample.

Characterization of Hsp70 proteins in bovine leukocytes induced by the temperature 41 degrees C.

Heat shock proteins (Hsp) are the group of proteins observed in both prokaryotic and eukaryotic cell types. Hsp synthesis takes place in response to many environmental conditions, including ultraviolet radiation, heavy metal ions, hypoxia and toxic agents. Many authors have suggested that Hsp can be used in immunoprophylaxis, yet Hsp70 proteins expressed in bovine leukocytes have not been fully characterized. Hence the aim of this study was to evaluate the expression of Hsp70 proteins in bovine leukocytes exposed to temp. 41degrees C. The material for the study consisted of bovine white blood cells incubated at 41 degrees C for 2 hours. SDS-Page electrophoresis, Western blotting, and two-dimensional electrophoresis (2D) were performed to estimate the proteins obtained. The results of the study confirmed the influence of the temperature of 41 degrees C on induction of Hsp70 in bovine leukocytes. These proteins were mainly localized within molecular mass 70kDa. Some of the proteins with molecular mass from 20 to 50 kDa also showed positive reactions in Western blotting with anti-Hsp70 antibodies. Analysis of 2D electrophoresis showed a change in the localization of these proteins in the pH gradient. It can be postulated that analysis of Hsp70 expression in bovine leukocytes can be a very helpful marker for evaluating an organism's adaptation to environmental heat stress. The proteins obtained also showed immunological reactivity with rabbit antibodies in Western blotting reactions, indicating that they can be used as protective factors in the pathogenesis of many diseases.

Comparison of immunogenic properties of native and inactivated Mannheimia haemolytica Lkt in calves.

The aim of this study was to compare the immunostimulatory properties of Lkt of M. haemolytica inactivated by formaldehyde and glutaraldehyde and to evaluate the neutralizing properties of anti-Lkt antibodies. The experiment was conducted on 20 Black-and-White Lowland calves of 100 kg body weight, assigned to 4 experimental groups. The animals were given subcutaneous vaccine injections with native Lkt, Lkt inactivated by formaldehyde or Lkt inactivated by glutaraldehyde. The anti-Lkt antibody titres were measured using an enzyme-linked immunosorbent assay (ELISA), based on absorbance of the sera obtained from the animals immunized with the different forms of Lkt. The protective effects of the antibodies present in the sera isolated from the vaccinated animals were estimated using an MTT assay. Analysis of the ELISA absorbance values in the sera from calves in the vaccinated groups did not show any significant differences between the groups. The highest increase in absorbance of sera was observed in calves from the group that received formaldehyde-inactivated Lkt. In the case of calves immunized with native Lkt, the absorbance values were lower than in the group immunized with Lkt inactivated by formaldehyde. The lowest absorbance values were observed in sera obtained from calves vaccinated with Lkt inactivated by glutaraldehyde. Analysis of the MTT assay results revealed the greatest Lkt-neutralizing properties of antibodies in the sera of calves immunized with two doses of a vaccine containing native Lkt and Lkt inactivated with formaldehyde.

Use of oromucosally administered interferon-alpha in the prevention and treatment of animal diseases.

Interferon-alpha (IFN-alpha) is well known as a clinically effective antiviral and antineoplastic therapeutic agent. It has also been shown to have immunoregulatory properties. IFN-alpha stimulates a cell-mediated innate immune response and then participates in the transition of the initial host innate response to an effective adaptive immune response. IFN-alpha is produced in small quantities in nasal secretions during viral infections, prompting many authors to suggest that low-dose oromucosal administration of IFN-alpha effectively mimics nature. Moreover, the injectable high-dose interferon therapy currently approved for various human disorders causes numerous side effects. By contrast, oromucosal administration of IFN-alpha is not associated with toxic effects. Another distinct advantage is ease of administration: the IFN can be dissolved in drinking water or administered by nebulization to the oral or nasal cavity. This review describes the current state of knowledge concerning orally administered IFN-alpha, of both human and animal origin, as a prophylactic or therapeutic agent in veterinary medicine. We present the effects of IFN-alpha in such animals as cattle, pigs, horses, cats, dogs and chickens, and attempt to explain its mechanism of action following oromucosal administration. It is hoped that this review of the medical literature on the use of IFN-alpha in animals will give practitioners a better understanding of the challenges and benefits of using this interesting cytokine in clinical practice.

Evaluation of Mannheimia haemolytica leukotoxin prepared in nonsupplemented and BSA or FBS supplemented RPMI 1640 medium.

Mannheimia haemolytica leukotoxin (Lkt) is the major factor that contributes to lung injury in bovine pneumonic pasteurellosis. Supernatant preparations containing Lkt produced by M. haemolytica serotype 1, grown in RPMI 1640 medium supplemented with BSA or FBS and without supplements were evaluated during this study. Analysis of obtained Lkt showed presence of 105 kDa antigen (SDS-PAGE electrophoresis). The obtained bacterial protein fraction estimated as Lkt was detected by Western blotting with mouse monoclonal (Mab 605 and Mab 601) anti-Lkt antibodies. No significant differences were found in obtained leukotoxin between wildtype and reference M. haemolytica strains. Our studies showed that growth in media supplemented with BSA or FBS had no significant influence on leukotoxin production. When BSA or FBS supplements were used, additional protein fractions in electrophoregrams SDS-PAGE were observed. These protein bands did not react with Mab 605 and/or Mab 601 in Western blotting analysis. Lkt immunogenicity was detected by immunoblotting with sera from Lkt immunized rabbits and calves.

The influence of transport stress on the humoral immunological response of calves induced by Mannheimia haemolytica leukotoxin.

In the contemporary systems of cattle production, transport stress is the most essential poly-etiological factor responsible for inducing unfavourable reactions in the animals. The main reason for this phenomenon is the immunosuppressive effect of steroid hormones on cellular and humoral protective mechanisms. The purpose of the present study was to establish the relationship between the cortisol concentration as an indicator of the stress reaction occuring directly after the transportation of calves and the specific humoral immune response to the leukotoxin (Lkt) antigen produced by the M. haemolytica strain. The experiment was carried out on 19 clinically healthy calves, weighing about 100 kg and transported by track for about 2 hours. After the delivery of the animals for feeding to the traditional cattle-house, the calves were immunized s.c.: group I with 1 ml Lkt (in conc.--10 microg/ml) with 1 ml of adjuvant on the 1st and 14th day after the transportation, group II with the same Lkt doses on the 3rd and 16th day after the transportation. The animals of the control group were vaccinated on the 1st and 14th day after the transportation with the twice diluted adjuvant. In examined sera the cortisol concentrations and the level of Lkt antibodies were measured by ELISA test. The cytotoxin neutralizing (CN) antibody level (cytotoxity assay) was determined with a simple visual assay. The study revealed, significant differences (P < or = 0.01) in serum cortisol levels between the control and experimental animals. The analysis of the absorbance of the sera in both groups immunized with Lkt showed substantial differences (P < or = 0.05) from the 6th through to 22nd day of the experiment compared with the control group. The analysis of the results of CN antibody titers showed no differences between the sera from group I and II. Based on the results obtained in this experiment it can be assumed, that a short transportation stress has no important influence on the level of specific humoral anti-Lkt response.

Characterization of Escherichia coli strains associated with canine pyometra.

The purpose of this study was to characterize E. coli isolates from canine pyometra which were isolated in pure culture. The E. coli strains were obtained in 128 cases, from 143 animals which were submitted to ovariohisterectomy. Biochemical analysis of all strains examined was possible on separation of 10 primary biotypes. The majority of the strains (87.5%) belonged to biotype 9, 1, 13 and 15. Dulcitol was fermented by 93% of all isolates. Haemolysin and colicin production was found in 53.9% and 26.6% of the strains, respectively. Approximately 37% of strains expressed resistance to two or more antibacterial agents. No plasmid was detected in 4.6% of the isolates. Plasmid profiles of all plasmid-containing isolates revealed plasmid bands corresponding to molecular weight ranging from 1 kb to 160 kb. Many of the strains examined had a single plasmid of 110 kb (46.1%), or two plasmids 110:65 kb (18.8%). Both plasmids appearing alone or in combination with other plasmids were detected in 90.1% of isolates with plasmid content. It was established that among haemolytic, colicinogenic and motile strains, the presence of both plasmids was 91.3, 94.1 and 91.4%, respectively. The appearance of both plasmids among dulcitol-positive and raffinose-negative strains was estimated at 88.2 and 88.3%, respectively. In a group of colicinogenic strains the presence of a single plasmid of molecular weight 110 kb was estimated at 5.9%. When both plasmids were present (profile 110:65), the percentage of these strains was 70.6%.

[Bipolar hysteroscopic electrosurgery done with Versapoint system: comparison to operations done with monopolar electrodes, own experience]. / Dwubiegunowa elektrochirurgia histeroskopowa przy zastosowaniu systemu VERSAPOINT--porównanie z zabiegami wykonywanymi elektrodami jednobiegunowymi, doswiadczenia wlasne.

The aim of the paper was to compare properties of mono- and bipolar electrodes in histeroscopic surgery. The bipolar electrodes allow enriching better hemostasis, thanks of the vaporisation effect it takes less time to remove submucosal myomas. Versapoint is an alternative tool for the laser. Use of physiologic saline as the medium for uterine cavity distension is safer for patients during long-lasting histeroscopic operations.

[Renoscintigraphy applied with etylenedicysteine labeled with technetium 99m and SPECT technique as a method of examining kidneys in children with vesicoureteric reflux]. / Renoscyntygrafia z zastosowaniem etylenodicysteiny znakowanej technetem 99m oraz technika SPECT jako metody badania nerek u dzieci z rozpoznanymi wstecznymi odplywami pechorzowo-moczowodowymi.

Vesicoureteric reflux (VUR) can lead to renal parenchymal damage. Renal scarring is an important cause of chronic renal failure and hypertension in children. The significance of possible effects determines the necessity of early diagnosis of urinary tract pathology. The aim of the paper was to evaluate the morphology and function of kidneys with VUR using selected radioisotope techniques, and to compare the sensitivity of planar technique and single-photon emission computed tomography (SPECT) technique in detection of renal scarring. In 45 children with VUR the following test were performed: ultrasonography, renoscintigraphy with technetium-99m-ethylenedicysteine (EC-Tc-99m) and technetium-99m-dimercaptosuccinic acid (DMSA-Tc-99m) scintigraphy with planar and SPECT mode. Stage of VUR correlates with stage of cortical lesions estimated as a value of effective renal plasma flow (ERPF) in kidney, as well as scarring intensity in static scintigraphy. The use of SPECT increases sensitivity of examination for detection small, single scars. It seems that SPECT should be used more frequently in children in the group of scarring risk. That would allow for earlier diagnosis of renal scarring, enabling efficient treatment. Due to the correlation between ERPF and parameters obtained in DMSA scintigraphy, renoscintigraphy with EC-Tc-99m may be applied to monitor the progress of renal scarring.

[The influence of cumulative dexamethasone, promethazine and dextran 70 used as protection against intraperitoneal adhesions on selected parameters of humoral immunity in women operated on for infertility]. / Wplyw zapobiegajacego zrostom sródotrzewnowym lacznego podawania deksametazonu, prometazyny i dekstranu 70 na wybrane parametry odpornosci humoralnej u kobiet operowanych z powodu nieplodnosci mechanicznej.

One of the most frequent reasons of the intraperitoneal adhesions formation in young women is pelvic inflammatory disease and surgical operations of pelvic organs. The element of more complex activities against adhesion creation is to limit an inflammatory reaction in the area of operation by intraperitoneal and systemic pharmacotherapy. The system of activities put into practice in the Clinic of Gynaecology includes in the field of pharmacological prophylaxis a systemic and intraperitoneal use of dexamethasone and promethazine before, during and after the operation, in total doses respectively: 250 mg and 300 mg, as well as intraperitoneal dosing of 500 ml of dextran 70, just before closing the peritoneal cavity. Verification of the results of these activities during laparoscopy or laparotomy shows its high effectiveness. For many years we did not observe any complications of any wound healing after the operations. Due to existing reports about possible complications in the postoperation period, which are probably caused by the use of the above-mentioned pharmaceutics, I decided to investigate how dexamethasone, promethazine and dextran 70 influence the selected parameters of humoral resistance in women who were operated on due to mechanical infertility. The investigated group consisted of 30 women, while the control group comprised 10 women who had gynaecological operations of similar extent. The venal blood samples were collected before the operation and on the third, fifth and twenty first day after the operation. The levels of selected acute phase proteins were checked and the character as well as the dynamics of value changes were monitored in both groups. Because of great variability between the proteins the fractions were picked out focusing on the differences in dynamics of metabolism, half-life time and biological functions. Following proteins were investigated: C-reactive protein, alfa1-acid glycoprotein, haptoglobin, alfa1-antitripsin, ceruloplazmin, alpha2-macroglobin, hemopexin, immunoglobulins A, G, M, albumins, transferrin, alpha2-HS-glycoprotein. The analysis of the study results has indicated that the application of dexamethasone, promethazine and dextran 70 only exerts modulating influence on the course of acute phase reaction induced by the operations. In the group having been studied earlier (in 3rd postoperative 24-hours) significant changes were revealed concerning the level of particular proteins in accordance with their physiological character (increase or decrease of level) in the control group such changes appeared later (about 5th 24-hours) and were characterized by markedly smaller amplitude and dynamics (Tab. 1-2, Fig. 1-15). The levels of proteins defined prior to operation were reached faster by patients of the study group (up to 21st 24-hours after operation). It has been disclosed that the use of composed antiadhesion therapy with dexamethasone, promethazine and dextran 70 accelerates, intensifies and simultaneously restricts the duration of transformations defined as acute phase reaction, which result in reducing the number of intraperitoneal adhesions. This therapy does not impair the mechanisms of humoral immunity in patients operated on.

[Evaluating the possibility of performing diagnostic and operative hysteroscopy depending on the type of hysteroscopy and agent used for filling the uterine cavity]. / Ocena mozliwosci wykonania diagnostycznych i operacyjnych hysteroskopii w zaleznosci od typu hysteroskopu i zastosowanego srodka wypelniajacego jame macicy.

The aim of the paper is to present own experiences concerning diagnostic and operative hysteroscopy performed with 4 types of hysteroscopes. Flexible hysteroscope HYP 1T, rigid hysteroscope 5 mm in diameter with single flow, operative hysteroscope 8 mm in diameter with constant flow, hysteroresectoscope 9 mm in diameter with constant flow produced by Olympus; for filling the uterine cavity CO2 0.2% NaCl, 5% glucose, Purisol, Hyskon were implemented. In even case the degree of hardship of the procedure, traumatism for the patient, degree of operated-on patient's safety during the procedure, possibility of applied in the presence of bleeding, price of instrument and agents applied for filling the uterine cavity were estimated. It has been found out that the use of rigid hysteroscope 5 mm in diameter and operative hysteroscope 8 mm in diameter with constant flow and utilization of CO2 as well as Purisol and Hyskon solutions provide possibility of extensive diagnosis and therapy.

[Developmental anomalies of the uterus in a population of 3000 women with various causes of infertility]. / Wady rozwojowe macicy w zbiorowosci 3000 kobiet z rózna przyczyna nieplodnosci.

In the years from 1979 to 1991 a population of 3000 women with various causes of infertility was found to have in 13.34% developmental anomalies of uteri. As many as 9.27% consisted of saddle uteri, 2.67% bicornous, in 0.37% there was partial, and in 0.13% complete septum, in 0.57% unicornous and in 0.33% double. The diagnosis of developmental anomalies involving uterus was based mainly on HSG, laparoscopic and hysteroscopic examinations.

[Effect of hydrotubation on the anatomical state of oviducts in women with mechanical infertility]. / Wplyw hydrotubacji na stan anatomiczny jajowodów u kobiet z nieplodnoscia mechaniczna.

The aim of the paper was to evaluate the anatomical state of uterine adnexa in women operated on due to mechanical infertility. Hydrotubation were previously applied in 31 women stemming from various centres in the country, while in 31 such a treatment was not performed at all. The mentioned groups of women were encumbered, to a similar degree, with a risk factor of infertility, except for the duration of sterility, which in those treated by hydrotubation lasted on the average 2 years longer. From 5 to 50 hydrotubations were carried out, most frequently in series of 5 procedures. It was reported by the women that 8 of them after hydrotubation experienced hypogastric pain persisting for some days, and in 6 there was acute adnexitis. Destructive changes in uterine adnexa, being estimated during the reconstructive operation, were decidedly more advanced in women treated by hydrotubation. That was expressed mainly by frequent appearance of lytic adhesions as well as by more advanced fibrosis of oviducts, particularly that of endosalpinx. Unchanged oviducts, after their release from adhesions, also appeared less frequently.