Detection of Pseudomonas aeruginosa in water samples using a novel synthetic medium and impedimetric technology.

AIMS: To investigate whether the use of a novel synthetic medium in conjunction with impedimetric technology could provide a rapid and automated detection of Pseudomonas aeruginosa in water samples. METHODS AND RESULTS: A selective synthetic medium (Z-broth) in which the only carbon and nitrogen source is acetamide was applied in direct impedimetric examination for the selective isolation of P. aeruginosa. A total of 1036 tap-water, well-water, swimming-pool water and dialysis water samples were investigated, and any P. aeruginosa contamination was detected in 7-24 h. Neither false-negative nor false-positive results were observed. CONCLUSIONS: The results of the present evaluation demonstrate that impedance measurement with the use of Z-broth is suitable for the rapid and automatic detection of P. aeruginosa in water. SIGNIFICANCE AND IMPACT OF THE STUDY: The main advantages of the method: 240 samples can be examined in one step, the procedure is fully automated, the results are obtained quickly and the labour and media requirements are low.

Radiodiagnostic examination of the swimbladder of some fish species.

Radiodiagnostic methods have not been used previously for studying the anatomy and diseases of the swimbladder of freshwater fish species. In this study, the radiographic anatomy of the swimbladder and species-related differences in swimbladder structure were studied on plain radiographs taken of 12 Hungarian fish species of major economic importance. Changes observed by radiography were also studied by conventional parasitological methods. The radiodiagnostic method reported here appears to be a useful complement to diagnostic examinations that have been based merely on dissection so far. It enables evaluation of the pathological lesions in live condition, without causing damage to the fish.

Plasminogen activator activity in tears after excimer laser photorefractive keratectomy.

PURPOSE: To quantify changes of plasminogen activator activity in tear fluid during corneal re-epithelialization after excimer laser photorefractive keratectomy (PRK). METHODS: Tear samples were collected with glass capillaries from 77 eyes of 42 patients immediately before and immediately after PRK treatment and on postoperative days 3 and 5. In 20 patients, the contralateral eye was similarly sampled to serve as control. Plasminogen activator activity in the tear samples was measured by a spectrophotometric method using human plasminogen and chromogenic peptide substrate, D-valyl-L-leucyl-L-lysine-p-nitroanilide (S-2251). RESULTS: In tears of all eyes that underwent PRK, the plasminogen activator activities were lower immediately after PRK than were the preoperative values. For patient eyes with normal wound healing, tear plasminogen activator activities were significantly elevated above the preoperative level on the third postoperative day and then returned to the preoperative level by the fifth postoperative day. In contrast, tear plasminogen activator activities remained low through the third postoperative day in all (six) eyes in which haze developed after 3 to 6 months. The contralateral control eyes showed no appreciable change in plasminogen activator activity over the 5-day period. CONCLUSIONS: Plasminogen activator activity levels measured in tears of excimer laser PRK-treated eyes may serve as a predictor of wound healing. Extended low levels of plasminogen activator activity through the third postoperative day correlate with the development of corneal healing abnormalities (haze). The low plasminogen activator activity could be not only an accompanying sign but also a cause of defective corneal wound healing.

Occurrence of acute paralysis virus of the honey bee (Apis mellifera) in a Hungarian apiary infested with the parasitic mite Varroa jacobsoni.

Viruses of the honey bee have been known for a long time; however, recently the attention of scientists and apiculturalists has turned towards the relationship between these viruses and the parasitic mite Varroa jacobsoni. Although clinical symptoms indicated the presence of some of the viruses of bees in Hungary, none have previously been isolated or identified. During July unusual adult bee and brood mortality was observed in some colonies of an apiary in Budapest known to be infested with Varroa jacobsoni. Large amounts of acute paralysis virus (APV) were detected serologically in healthy honey bee pupae killed by the injection of a bacteria-free extract of diseased adult bees. Crystalline arrays of 30 nm particles were seen in ultrathin sections of the tissues of injected pupae and naturally infected adult bees. In spite of the application of acaricide treatments the bee population in several colonies had collapsed by the end of summer and the apiary suffered severe wintering losses.

Radiodiagnostic method for studying swimbladder inflammation caused by Anguillicola crassus (Nematoda: Dracunculoidea).

The nematode Anguillicola crassus appeared in Europe about 15 yr ago, and has subsequently become recognised as a cause of major pathological lesions in the swimbladder of the European eel Anguilla anguilla. The radiographic method reported in this paper showed be a useful complement to diagnostic methods that have so far been based exclusively on fish dissection. Using this method, the infection of the swimbladder and the severity of its pathological changes can be assessed in a reliable manner without causing damage to the fish. By analysing radiographs of 45 eels from Lake Balaton, 5 stages of swimbladder lesions were distinguished. The method enables the examiner to draw conclusions on the air, worm and exudate content of the swimbladder; however, data on thickening of the swimbladder wall can be obtained only indirectly. The radiograph also provides information on the air and worm content of the pneumatic duct. Results obtained by radiography showed good agreement with dissection findings. Radiographic results are compared with pathological findings obtained from fish dissections separately for each severity grade of infection.

Interaction of chicken anaemia virus and Cryptosporidium baileyi in experimentally infected chickens.

The natural occurrence of concomitant chicken anaemia virus (CAV) and Cryptosporidium baileyi infection was described earlier. In this experiment, 1-day-old chickens were infected with CAV alone (anaemia virus infected, AI) or followed by inoculation with 8 x 10(5) C. baileyi oocysts orally at 1 wk of age (anaemia virus and Cryptosporidium infected, ACI). Another group of chickens received the same dose of C. baileyi oocysts without previous virus infection (Cryptosporidium infected, CI), and two groups of uninfected chickens served as controls. Except one group (uninfected control, UC), all groups -- including the other control group (challenged control, CC) -- were challenged with an oral inoculum of 8 x 10(5) C. baileyi oocysts at the age of 4 wk. Haematological, serological, immunohistochemical and pathological findings confirmed the effect of the virus agent. The individual C. baileyi oocyst shedding did not show significant difference between group CI and ACI, however, after challenge infection the AI chickens shed approximately three times more C. baileyi oocysts than those in group CC. Mortality and the percentage of birds that developed anaemia was significantly higher among ACI than AI chickens, while haematocrit values at 2 wk of age and relative bursal weights at 4 wk of age were moderately lower in the ACI group. The results presented here suggest that concurrent CAV infection increases the reproductive potential of C. baileyi in chickens, and both pathogens have synergistic effect on each other.

Attempts to immunize chickens against Cryptosporidium baileyi with C. parvum oocysts and Paracox vaccine.

To study the possibility of immunization against Cryptosporidium baileyi Current, Upton et Haynes, 1986 with the attenuated anticoccidial vaccine, Paracox and oocysts of C. parvum Tyzzer, 1912, chickens were inoculated orally with either 3 x 10(3) vaccine oocysts or 8 x 10(5) C. baileyi or C. parvum oocysts at 1 week of age. The inoculation with Paracox vaccine and C. parvum oocysts was repeated at 2 and 3 weeks of age. Uninfected birds served as controls. All animals with the exception of one uninfected group were challenged orally with either 8 x 10(5) C. baileyi or 3 x 10(5) Eimeria tenella Railliet et Lucet, 1891 oocysts at 4 weeks of age. Sera were collected at 4 weeks of age, and were examined by ELISA using C. baileyi antigens. Birds inoculated with C. parvum oocysts did not shed C. parvum oocysts in their faeces, but anticryptosporidial antibodies could be detected in the sera. The total oocyst output of C. parvum inoculated chickens was 17% of that of previously uninfected birds after the oral challenge with C. baileyi. Considering that antibodies play no or only a minor role in resistance to C. baileyi, these results suggest that inoculation of chickens with C. parvum oocysts stimulated also cellular immune response. Based on the relative body weight gain, faecal scores, oocyst output, mortality, and caecal lesions in the birds immunized with Paracox vaccine and challenged with E. tenella, the vaccination induced only a moderate protection against the reinfection. The results of cross-immunization of chickens with Eimeria spp. and C. baileyi suggest that attenuated anti-eimerian vaccines do not induce any protection against cryptosporidial infection.

Scleral reinforcement in progressive myopia and intraoperative ultrasound control of the cadaver fascia lata strip.

Scleral reinforcement (sustentaculum sclerae) is one of the operations against myopia. During the progression of myopia the eye grows. The chorioretinal layer can only moderately follow the expansion of the sclera, and mostly this is the cause of the complications that can cause permanent visual acuity decrease. The aim of the operation is to strengthen the posterior part of the sclera by implantation of a cadaver fascia lata strip, that slows down or stops the expansion of the eyeball, and this way prevents the development of complications. At our Department we perform the Snyder-Thompson technique. Between 1984 and 1994 343 operations were performed at our Department. We followed up the changes of the axial lengths by ultrasound (A-mode) examinations. The axial length decreased in 43.7% remained unchanged in 22.2% and increased in 34.1% of the cases. We controlled the location of the strip intra- and postoperatively with B-mode ultrasound. Nowadays scleral reinforcement is still the only possibility to stop or slow down the expansion of the eyeball in cases of progressive high myopia.

Excimer laser photorefractive keratectomy with different ablation zones.

In this study we would like to introduce the excimer laser, and to demonstrate our results and complications by using different ablation zones during photorefractive keratectomy (PRK) in the correction of myopia and astigmatismus. In 1996 we performed photorefractive keratectomy on 100 myopic eyes of 52 patients (28 females, 24 males). Mean age was 26.21 years (ranged from 19 to 54 years). The preoperative refraction ranged from -1.0 D to -18.0 Diopters. The diameter of the ablation zones were between 5 and 6.5 mm. We evaluated the results and the complications of the surgeries of 100 eyes which were performed with Schwind keratom F excimer laser. After 2 days, 1 week, 1 month, 3 months, and 6 months postoperatively we tested the best uncorrected and corrected visual acuities, and performed intraocular pressure measurement, slit lamp examination as well as corneal topography. The postoperative refractions were between +/- 0.5 to +/- 1.0 Diopters. After six months postoperatively the slit lamp examination showed that 80% of the patients had no corneal haze while 20% had stage I (Hanna) corneal haze. The smaller the diameter of the ablation zone was, the more pronounced the corneal haze and the night-glare were. The photorefractive excimer laser keratectomy is judged to be a safe method, although it might have some side-effects. The different ablation zones of this treatment means an important modification, that not only allows the method to meet the individual requirements, but reduces the chance of the complications as well. Based on the authors' experiences PRK for moderate myopia with large diameter ablation zones appears more predictable than than with smaller ablation zone diameters.

In ophthalmology new possibilities for the use Nd:YAG laser.

Results of the technical development in laser technology created new possibilities in ophthalmic surgery, too. The easy access to various lasers offers an extraordinary opportunity for eye surgeons. At the Department of Ophthalmology of the University Medical School of Debrecen we performed 1400 laser treatments in 1996. Out of that 1100 were retinal Argon laser coagulations because of diabetic retinopathy, 200 YAG laser capsulotomies because of secondary cataract, and 30 laser iridotomies because of glaucoma. Additional 70 laser treatments were performed for special indications. During the laser treatment of the retina and the choroid we use Argon gas lasers, whereas the Nd:YAG laser used in the anterior segment of the eye is a solid-state laser. Our pulse mode modus synchronized Nd:YAG apparatus can produce megawatts of energy in nanoseconds. Authors give an account of the Nd:YAG lasers' possibilities in therapy (capsulotomy, iridotomy), present the most frequent indications of treatments in 1996, and compare it with international literature data. Authors demonstrate the newest principles of the Nd:YAG laser treatment of secondary membranes that are occasionally formed in the anterior chamber especially in the area of the pupil following cataract surgery.

Attempts to analyse Anguillicola crassus infection and the humoral host response in eels (Anguilla anguilla) of Lake Balaton, Hungary.

Since the introduction of Anguillicola crassus into Europe, anguillicolosis has been a considerable problem in several countries. From 1991, periodical eel mortality occurred in Lake Balaton, Hungary. However, eels with a worm burden of 20 to 50 parasites did not show severe swimbladder lesions, which observation cast doubts on the primary aetiological role of the parasite in the eel kill. In order to study the pathology of the infection, from the spring of 1996 until October of the same year, 51 eels were collected from two regions of Lake Balaton and examined for swimbladder changes. To detect humoral antibodies, an enzyme-linked immunosorbent assay (ELISA) was performed, using cuticular-oesophageal worm antigen. The results of the test show the applicability of the method. However, no direct correlation was found between antibody levels or the intensity of infection and the swimbladder lesions. The low level of specific antibodies and the increasing severity of swimbladder changes in the autumn suggest that parasite-induced immunity is insufficient to prevent reinfection.

Effect of F-2 and T-2 fusariotoxins on experimental Cryptosporidium baileyi infection in chickens.

The course of Cryptosporidium baileyi infection in chickens fed with different doses of fusariotoxins was compared with that of control groups. F-2 toxin levels of 0.187-1.5 mg kg-1 and T-2 toxin levels of 0.187-6.0 mg kg-1 were investigated. The experimental animals were orally infected with 6 x 10(5) C. baileyi oocysts at 1 week of age. Total daily oocyst output was monitored by a quantitative method. Acquired immunity was tested at the age of 4 weeks, by ELISA and by a challenge infection with an equal number of oocysts, upon recovery from the primary infection. The results show that in chickens kept on the lower doses of F-2 and T-2 toxins, the parasite infection ran a similar course to that in the control groups, and the animals became resistant to re-infection. However, when higher doses (2.0-6.0 mg kg-1) of T-2 toxin were used, a depression of weight gain was observed with some other physiological parameters (PCV, weight of bursa, weight of thymus, skin thickness in PHA-P skin test) also indicating toxic effect and, simultaneously, the oocyst output decreased significantly and the patent period was slightly prolonged. Although certain modifications of the immune response could be revealed, the chickens became resistant to re-infection. Only early (1 week of age) parasite infection and 6 mg kg-1 T-2 toxin in the feed significantly depressed body weight gain and immunity.

Attempts to immunize chickens with Cryptosporidium baileyi oocyst extract.

In order to study the possibility of immunization against Cryptosporidium baileyi with extracted crude antigen, Arbor Acres chickens were injected intramuscularly with 80 micrograms of C. baileyi oocyst-derived proteins (uninfected immunized, UI) or inoculated orally with 8 x 10(5) viable C. baileyi oocysts (infected control, IC) at 1 wk of age. The immunization was repeated in the UI group at 2 wk of age. Uninfected (UC) birds served as controls. All animals in UI, IC, and UC groups were challenged orally with 8 x 10(5) C. baileyi oocysts at the age of 4 wk. Blood samples were collected when birds were 4 and 6 wk of age, and sera were examined by enzyme-linked immunosorbent assay for the presence of antibodies against C. baileyi. Total oocyst output of UI chickens was about 60% of that of UC birds after challenge, and the prepatent and patent periods were nearly identical in the latter 2 groups. In contrast, IC birds developed complete resistance to challenge infections. These results suggest that immunization with the oocyst extract of C. baileyi may confer some degree of protection against oral challenge; however, the protection is less effective than that induced by primary oral infection. The lack of significant difference between the antibody responses of IC and UI animals to C. baileyi at 2 wk of age suggests that serum antibodies play little role in acquired resistance to challenge infection.

Effects of bursectomy and thymectomy on the development of resistance to Cryptosporidium baileyi in chickens.

Comparisons were made between the course of infection with Cryptosporidium baileyi in normal chickens and in chickens with functional deficiencies in either B-lymphocytes (bursectomized) or T-lymphocytes (thymectomized). Bursectomy did not influence the acquired immune response to C. baileyi infection as measured by the oocyst excretion. However, the total oocyst output of bursectomized birds was less than a quarter of that of the controls due to the reduced site for multiplication of the parasite. The total oocyst output of thymectomized chickens was more than 2 times higher and the patency was 2 times longer as compared with control animals. Moreover, thymectomized birds failed to acquire resistance to the challenge infection. These findings attest to the primary role of cell-mediated immunity in the expression of resistance to C. baileyi as opposed to the antibody-mediated mechanisms.

Potentiation of ionophorous anticoccidials with duokvin: battery trials against Cryptosporidium baileyi in chickens.

Three battery tests, employing 444 Arbor Acres chickens, were conducted to study the anticryptosporidial efficacy of the polyether ionophorous anticoccidials alone and in combination with a new dihydroquinoline antioxidant, duokvin. The following efficacy percentages for the ionophores at the recommended level were established on the basis of oocyst output: lasalocid 45, monensin 37, semduramicin 29, narasin 23, and maduramicin and salinomycin 21. The efficacy of the ionophores, with the exception of lasalocid, could be increased to 36-88% by combination with duokvin. However, these combinations resulted in a significant weight gain reduction due to toxic interaction. Efficacy showed a strong negative correlation with body weight gain and bursa weight, suggesting an indirect mode of action against the parasite. It has been concluded that neither ionophorous anticoccidials nor their combinations with duokvin can be recommended for chemoprophylaxis against avian cryptosporidiosis.

Age-dependent resistance to Cryptosporidium baileyi infection in chickens.

The age-dependent aspects of resistance to Cryptosporidium baileyi were studied in chickens orally inoculated with C. baileyi oocysts at 1 and 9 wk of age. The prepatent period was significantly shorter and the patency was significantly longer in younger birds. Chickens infected at 1 wk of age excreted 3 times the number of oocysts excreted by those inoculated at 9 wk of age. There was a good correlation between the length of the patent period and the total oocyst output of chickens. These results indicate that both innate and acquired resistance to C. baileyi are age-related. The numbers of animals needed per experimental group to estimate the length of the prepatent period, the patency, and the total oocyst output were assessed. Relatively small numbers of animals are required for the estimation of the mean length of the prepatent period, in contrast with the large numbers of birds needed for the estimation of the mean length of the patent period or that of the total oocyst excretion.

Quantitative method to assess Cryptosporidium oocyst shedding in the chicken model.

An unsophisticated and fairly sensitive quantitative method and its grounds are described, which proved useful to assess Cryptosporidium oocyst shedding in the chicken model. The method is based on the rather slow sedimentation of oocysts. A threshold of sensitivity ranging between 5,000 and 10,000 oocysts/g feces was established for this technique. There was good agreement between triplicate assays over a wide range of oocyst concentrations.

Concurrent cryptosporidiosis and chicken anaemia virus infection in broiler chickens.

Concurrent infection with Cryptosporidium baileyi and chicken anaemia virus (CAV) was observed in a flock of 8000 4-week-old broiler chickens. The birds, showing overt symptoms of stunted growth and 25% mortality from hatching to 4 weeks of age, harboured the protozoan in the epithelial cells of the bursa of Fabricius and the urodeal portion of the cloaca. This is the first report on an outbreak of avian cryptosporidiosis associated with CAV.