Genotyping of European Toxoplasma gondii strains by a new high-resolution next-generation sequencing-based method.

PURPOSE: A new high-resolution next-generation sequencing (NGS)-based method was established to type closely related European type II Toxoplasma gondii strains. METHODS: T. gondii field isolates were collected from different parts of Europe and assessed by whole genome sequencing (WGS). In comparison to ME49 (a type II reference strain), highly polymorphic regions (HPRs) were identified, showing a considerable number of single nucleotide polymorphisms (SNPs). After confirmation by Sanger sequencing, 18 HPRs were used to design a primer panel for multiplex PCR to establish a multilocus Ion AmpliSeq typing method. Toxoplasma gondii isolates and T. gondii present in clinical samples were typed with the new method. The sensitivity of the method was tested with serially diluted reference DNA samples. RESULTS: Among type II specimens, the method could differentiate the same number of haplotypes as the reference standard, microsatellite (MS) typing. Passages of the same isolates and specimens originating from abortion outbreaks were identified as identical. In addition, seven different genotypes, two atypical and two recombinant specimens were clearly distinguished from each other by the method. Furthermore, almost all SNPs detected by the Ion AmpliSeq method corresponded to those expected based on WGS. By testing serially diluted DNA samples, the method exhibited a similar analytical sensitivity as MS typing. CONCLUSION: The new method can distinguish different T. gondii genotypes and detect intra-genotype variability among European type II T. gondii strains. Furthermore, with WGS data additional target regions can be added to the method to potentially increase typing resolution.

A ring trial to harmonize Toxoplasma gondii microsatellite typing: comparative analysis of results and recommendations for optimization.

A ring trial among five European laboratories was organized to reach consistency in microsatellite (MS) typing of the zoonotic parasite Toxoplasma gondii. Three sample sets were circulated and analyzed by each laboratory following a previously published method that is based on fragment length polymorphism of 15 MS markers. The first sample set compared typing results in general and focused on effects of DNA concentration; the second sample set focused on the polymorphic fingerprinting markers that can differentiate T. gondii strains within the same archetypal lineage; and the third set focused on non-archetypal genotypes. Methodological variations between laboratories, including the software programs used to determine MS fragment length, were collated using a questionnaire. Overall, lineage-level typing results reached a high level of agreement, especially in samples with the highest DNA concentrations. However, laboratory-specific differences were observed for particular markers. Major median differences in fragment length, of up to 6 base pairs, were related to the fluorophore used to label fragment-specific primers. In addition, primer pairs with identical sequences obtained from different suppliers resulted in fragments of differing length. Furthermore, differences in the way the sequencing profiles were assessed and interpreted may have led to deviating results in fragment length determination. Harmonization of MS typing, for example, by using the same fluorophores or by numerical adjustments applied to the fragment-lengths determined, could improve the uniformity of the results across laboratories. This is the first interlaboratory comparison, providing guidelines (added as a supplement) for the optimization of this technique.

Topography of Spin Liquids on a Triangular Lattice.

Spin systems with frustrated anisotropic interactions are of significant interest due to possible exotic ground states. We have explored their phase diagram on a nearest-neighbor triangular lattice using the density-matrix renormalization group and mapped out the topography of the region that can harbor a spin liquid. We find that this spin-liquid phase is continuously connected to a previously discovered spin-liquid phase of the isotropic J_{1}-J_{2} model. The two limits show nearly identical spin correlations, making the case that their respective spin liquids are isomorphic to each other.

Disorder-Induced Mimicry of a Spin Liquid in YbMgGaO_{4}.

We suggest that a randomization of the pseudodipolar interaction in the spin-orbit-generated low-energy Hamiltonian of YbMgGaO_{4} due to an inhomogeneous charge environment from a natural mixing of Mg^{2+} and Ga^{3+} can give rise to orientational spin disorder and mimic a spin-liquid-like state. In the absence of such quenched disorder, 1/S and density matrix renormalization group calculations both show robust ordered states for the physically relevant phases of the model. Our scenario is consistent with the available experimental data, and further experiments are proposed to support it.

Chicken line-dependent mortality after experimental infection with three type IIxIII recombinant Toxoplasma gondii clones.

Three genetically different clones of Toxoplasma gondii, also different in mouse virulence, were studied by experimental infection in chickens. For the experiments, four chicken lines were used, which differed in phylogenetic origin and performance level: two white egg layer lines, one with high laying performance (WLA), one with low (R11) and two brown layer lines, also displaying high (BLA) and low (L68) egg number. Chickens were intraperitoneally infected with three different T. gondii isolates representing type IIxIII recombinant clones, i.e. showing both, type II- and type III-specific alleles. These clones (K119/2 2C10, B136/1 B6H6, K119/2 A7) had exhibited virulence differences in a mouse model. In chickens, a significantly higher mortality was observed in white layer lines, but not in brown layer lines, suggesting that differences in the phylogenetic background may influence the susceptibility of chickens for toxoplasmosis. In addition, antibody (IgY) levels varied in surviving chickens at 31 days post infection. While low to intermediate antibody levels were observed in white layers, intermediate to high levels were measured in brown layers. Infection with a T. gondii clone showing low chicken virulence resulted in higher antibody levels in all chicken lines compared to infection with T. gondii clones of intermediate or high chicken virulence. This was in agreement with the parasite load as determined by real-time PCR. Overall, results show that progeny resulting from natural sexual recombination of T. gondii clonal lineages, may differ in their virulence for mice and chickens.

Damped Topological Magnons in the Kagome-Lattice Ferromagnets.

We demonstrate that interactions can substantially undermine the free-particle description of magnons in ferromagnets on geometrically frustrated lattices. The anharmonic coupling, facilitated by the Dzyaloshinskii-Moriya interaction, and a highly degenerate two-magnon continuum yield a strong, nonperturbative damping of the high-energy magnon modes. We provide a detailed account of the effect for the S=1/2 ferromagnet on the kagome lattice and propose further experiments.

Besnoitia besnoiti infections activate primary bovine endothelial cells and promote PMN adhesion and NET formation under physiological flow condition.

Besnoitia besnoiti is an obligate intracellular and emerging coccidian parasite of cattle that mainly infects host endothelial cells during acute infection. We here analyzed early innate immune reactions of B. besnoiti-infected primary bovine umbilical vein endothelial cells (BUVEC). B. besnoiti infections significantly activated BUVEC since the gene transcripts of several adhesion molecules (P-selectin, intercellular adhesion molecule 1(ICAM-1)), chemokines (CXCL1, CXCL8, CCL5), and of COX-2 were significantly upregulated during in vitro infection. Overall, the highest upregulation of most transcripts was observed at 24 or 48 h post infection (p.i.). Enhanced adhesion molecule expression in infected host cells was confirmed by PMN adhesion assays being performed under physiological flow conditions revealing a significantly increased PMN adhesion on B. besnoiti-infected BUVEC layers at 24 h p.i. Furthermore, we were able to illustrate neutrophil extracellular traps (NETs) being released by PMN under physiological flow conditions after adhesion to B. besnoiti-infected BUVEC layers. The present study shows that B. besnoiti infections of primary BUVEC induce a cascade of pro-inflammatory reactions and triggers early innate immune responses.

Sarcocystis sinensis is the most prevalent thick-walled Sarcocystis species in beef on sale for consumers in Germany.

Bovines are intermediate hosts of Sarcocystis cruzi, Sarcocystis hirsuta, and Sarcocystis hominis, which use canids, felids, or primates as definitive hosts, respectively. Cattle represent also intermediate hosts of Sarcocystis sinensis, but the definitive hosts of this parasite are not yet known. Sarcocystosis in cattle is frequently asymptomatic. The infection is characterized by the presence of thin-walled (S. cruzi) or thick-walled muscle cysts or sarcocysts (S. hominis, S. sinensis, and S. hirsuta). Recent reports suggest high prevalence of the zoonotic S. hominis in beef in Europe. We therefore aimed at differentiating Sarcocystis spp. in beef offered to consumers in Germany using molecular and microscopical methods, focusing on those species producing thick-walled sarcocysts. A total of 257 beef samples were obtained from different butcheries and supermarkets in Germany and processed by conventional and multiplex real-time PCR. In addition, 130 of these samples were processed by light microscopy and in 24.6% thick-walled cysts were detected. Transmission electron microscopical analysis of six of these samples revealed an ultrastructural cyst wall pattern compatible with S. sinensis in five samples and with S. hominis in one sample. PCR-amplified 18S ribosomal DNA (rDNA) fragments of 28 individual thick-walled cysts were sequenced, and sequence identities of ≥98% with S. sinensis (n = 22), S. hominis (n = 5) and S. hirsuta (n = 1) were observed. Moreover, nine Sarcocystis sp. 18S rDNA full length gene sequences were obtained, five of S. sinensis, three of S. hominis, and one of S. hirsuta. Out of all samples (n = 257), 174 (67.7%) tested positive by conventional PCR and 179 (69.6%) by multiplex real-time PCR for Sarcocystis spp. Regarding individual species, 134 (52%), 95 (37%), 17 (6.6%), and 16 (6.2%) were positive for S. cruzi, S. sinensis, S. hirsuta, and S. hominis, respectively. In conclusion, S. sinensis is the most prevalent thick-walled Sarcocystis species in beef offered for consumption in Germany. Further studies are needed to identify the final host of S. sinensis as well as the potential role of this protozoan as a differential diagnosis to the zoonotic species S. hominis.

Isolation and molecular characterization of Toxoplasma gondii from captive slender-tailed meerkats (Suricata suricatta) with fatal toxoplasmosis in Argentina.

In this study, the diagnosis of fatal disseminated toxoplasmosis in three captive slender-tailed meerkats (Suricata suricatta) in the zoo of La Plata, Argentina and the invitro isolation and molecular characterization of Toxoplasma gondii are reported. The animals showed depression, dyspnea and hypothermia, and also ataxia in one case, and died within 1-5 days. The main histopathological lesions included interstitial pneumonia, non-suppurative inflammatory changes and focal necrosis in liver, spleen, kidney and brain. Tachyzoites or tissue cysts were present in lung, liver, spleen, brain, striated muscle, kidney, intestine and mesenteric lymph node sections, and stained strongly with T. gondii antiserum in immunohistochemical analysis. T. gondii was isolated in Swiss mice and in bovine monocytes cultures from tissues of one of the meerkats. The isolate was cryopreserved and it was named TG-Suricata-1. T. gondii DNA was demonstrated in tissues of all three animals and in tachyzoites isolated in cell cultures. The PCR-RFLP analysis of markers based in the loci 3'-SAG2, 5'-SAG2, BTUB, GRA6, SAG3, c22-8, L358, PK1, c29-2 and Apico of T. gondii produced patterns corresponding to the clonal type III. Type III strains of T. gondii possess no or only little virulence in the mouse model, however their association with virulence in other animal species is uncertain. In the present case, T. gondii of the clonal lineage III was responsible for fatal cases in S. suricatta. To our knowledge, this is the first report of isolation and genotyping of T. gondii from S. suricatta.

Validation of PCR-based protocols for the detection of Echinococcus multilocularis DNA in the final host using the Intestinal Scraping Technique as a reference.

Oral uptake of infectious Echinococcus multilocularis eggs shed by canids with their faeces may lead to development of alveolar echinococcosis in humans, which is clinically similar to a malignant infiltrative tumor and may be fatal if left untreated. E. multilocularis is therefore regarded as one of the most important and neglected metazoan parasites in the Northern hemisphere. The diagnosis of this tapeworm in the final host plays a key role in the epidemiology of E. multilocularis. The diagnostic performance of a magnetic-capture (MC) DNA extraction protocol in combination with a minor groove-binder real time PCR (MC-MGBqPCR) for the detection of E. multilocularis eggs was determined relative to a highly sensitive variant of the Intestinal Scraping Technique (IST) using faecal samples of foxes. In addition, we compared results obtained by MC-MGBqPCR with those of a previously validated protocol (QIAamp Fast DNA Stool Mini Kit (QT) combined with a TaqMan qPCR). Furthermore, a workflow using the NucleoMagVet DNA extraction kit (NM) in combination with MGBqPCR and TaqMan-qPCR was also included in the comparisons. To estimate the analytical sensitivity, phosphate-buffered saline and fox faecal samples were spiked with different numbers of eggs and tested in defined combinations of DNA extraction and PCR protocols. To assess the diagnostic sensitivity of the different workflows, samples were used that had been collected from the ampulla recti or the rectum of 120 foxes hunted in Brandenburg, Germany. The samples represented five IST categories formed according to the E. multilocularis worm burden of the foxes. For DNA extraction by MC or using two other commercial extraction kits, the supernatants obtained from 3 g of bead-beaten faecal samples were used. The extracted DNAs were then processed in the respective PCR protocols. The MC-MGBqPCR showed the highest diagnostic sensitivity (93%; 95% Confidence Interval (CI): 86-97%) relative to IST. The QT extraction protocol in combination with TaqMan-qPCR had the second highest sensitivity (89%; 95% CI: 80-94%), followed by NM with MGBqPCR (86%; 95% CI: 77-93%) in comparison to IST. The lowest diagnostic sensitivity was found for the NM combined with the TaqMan-qPCR protocol (72%; 95% CI: 62-82%). In conclusion, the MC-MGBqPCR seems to represent a suitable alternative to IST. However, applied to 3 g faecal samples, the less costly QT-TaqMan-qPCR workflow yielded a similar diagnostic sensitivity relative to IST. However, differences between these two workflows were not statistically significant.

Toxoplasma gondii infection and toxoplasmosis in farm animals: Risk factors and economic impact.

The protozoan parasite Toxoplasma gondii is a zoonotic parasite that can be transmitted from animals to humans. Felids, including domestic cats, are definitive hosts that can shed oocysts with their feces. In addition to infections that occur by accidental oral uptake of food or water contaminated with oocysts, it is assumed that a large proportion of affected humans may have become infected by consuming meat or other animal products that contained infective parasitic stages of T. gondii. Since farm animals represent a direct source of infection for humans, but also a possible reservoir for the parasite, it is important to control T. gondii infections in livestock. Moreover, T. gondii may also be pathogenic to livestock where it could be responsible for considerable economic losses in some regions and particular farming systems, e.g. in areas where the small ruminant industry is relevant. This review aims to summarize actual knowledge on the prevalence and effects of infections with T. gondii in the most important livestock species and on the effects of toxoplasmosis on livestock. It also provides an overview on potential risk factors favoring infections of livestock with T. gondii. Knowledge on potential risk factors is prerequisite to implement effective biosecurity measures on farms to prevent T. gondii infections. Risk factors identified by many studies are cat-related, but also those associated with a potential contamination of fodder or water, and with access to a potentially contaminated environment. Published information on the costs T. gondii infections cause in livestock production, is scarce. The most recent peer reviewed reports from Great Britain and Uruguay suggest annual cost of about 5-15 million US $ per country. Since these estimates are outdated, future studies are needed to estimate the present costs due to toxoplasmosis in livestock. Further, the fact that T. gondii infections in livestock may affect human health needs to be considered and the respective costs should also be estimated, but this is beyond the scope of this article.

Analysis of Toxoplasma gondii clonal type-specific antibody reactions in experimentally infected turkeys and chickens.

Due to their ground-feeding behaviour, free-ranging chickens and turkeys are exposed to oocysts and are good indicators of the presence of Toxoplasma gondii in the environment. In addition, poultry may become infected by ingestion of tissues of infected intermediate hosts such as small rodents. Free-ranging poultry are considered an important source of T. gondii infection in humans, especially in developing countries. Knowledge on T. gondii genotypes in infected animals and humans is important for understanding the epidemiology of T. gondii infections. The aim of the present study was to analyse the ability of experimentally infected turkeys and chickens to develop a T. gondii clonal type-specific antibody response (IgY) after i.v. inoculation with tachyzoites of three T. gondii clonal lineages, types I, II and III. A peptide microarray displaying a panel of 101 different synthetic peptides was used for serotyping. Peptide sequences were derived from polymorphic regions of 16 T. gondii proteins (GRA1, GRA3-7, SAG1, SAG2A, SAG3, SAG4, SRS1, SRS2, ROP1, NTPase I and NTPase III and BSR4). The array was probed with 120 sera from experimentally infected chickens and turkeys inoculated with different doses of T. gondii tachyzoites (104, 103 and 102) collected from isolates representative for T. gondii clonal types I (RH), II (ME49) or III (NED) and uninfected controls. After screening of the peptides with reference sera from chickens and turkeys, and evaluation of data by Receiver Operating Characteristics analysis, 41 and 40 peptides were identified that appeared suitable to detect type-specific reactions with sera collected at 2, 5, 7 and 9 weeks p.i. Selected peptides allowed the identification of T. gondii clonal types, until 9 week p.i., which the chickens or turkeys had been inoculated with. At 9 weeks p.i., a high proportion of the experimentally infected chickens (67% (12/18)) and turkeys (61% (11/18)) no longer reacted with the selected peptides. Serotyping of the infection in individual chickens or turkeys was only possible when the whole peptide panel was applied. Clonal type-specific antibody responses were dynamic in both poultry species and depended on the individual animal and the time after infection.

Toxoplasma gondii infections in chickens - performance of various antibody detection techniques in serum and meat juice relative to bioassay and DNA detection methods.

Chickens, especially if free-range, are frequently exposed to Toxoplasma gondii, and may represent an important reservoir for T. gondii. Poultry products may pose a risk to humans, when consumed undercooked. In addition, chickens are regarded as sensitive indicators for environmental contamination with T. gondii oocysts and have been used as sentinels. The aim of the present study was to determine the suitability of commonly used antibody detection methods, i.e. the modified agglutination test (MAT), IFAT and ELISA to detect T. gondii-infected chickens. Samples of experimentally and naturally infected chickens were used. The infection state of all chickens was determined by Magnetic-Capture (MC-) real-time PCR (RT PCR). Naturally exposed chickens were additionally examined by mouse bioassay and conventional RT PCR on acidic pepsin digests (PD-RT PCR). Blood serum and meat juice of various sources were tested for antibodies to T. gondii. In naturally infected chickens, there was substantial agreement between the mouse bioassay and MC-RT PCR or the mouse bioassay and conventional PD-RT PCR. PD-RT PCR was slightly more sensitive than MC-RT PCR, as all (26/26) bioassay-positive chickens also tested positive in at least one of the tissues tested (heart, drumstick). By MC-RT PCR, 92.3% (24/26) of the naturally infected bioassay-positive chickens were positive. The diagnostic sensitivity of MC-RT PCR was clearly related to the organ examined. Based on a quantitative assessment of the MC-RT PCR results in experimentally infected chickens, brain and heart tissues harbored an at least 100 times higher parasite concentration than breast, thigh or drumstick musculature. In naturally infected chickens, only three out of 24 birds, which were MC-RT PCR-positive in heart samples, also tested positive in drumstick musculature. Under experimental conditions, the agreement between MC-RT PCR and the serological techniques revealed 100% diagnostic sensitivity and specificity. Under field conditions, examinations of sera by ELISA, IFAT and MAT showed good performance in identifying chickens that were positive in either a mouse bioassay, MC-RT PCR, or PD-RT PCR as illustrated by diagnostic sensitivities of 87.5%, 87.5% and 65.2%, respectively, and diagnostic specificities of 86.2%, 82.8% and 100%, respectively. The examination of meat juice samples from breast, drumstick or heart musculature revealed similar or even better results in the ELISA. The results in the MAT with meat juice from breast musculature were less consistent than those of ELISA and IFAT because a number of negative chickens tested false-positive in the MAT. The MAT performed similar to ELISA and IFAT when applied to test meat juice samples collected from heart, thigh or drumstick musculature.

High seroprevalence of Toxoplasma gondii and probability of detecting tissue cysts in backyard laying hens compared with hens from large free-range farms.

Serological assays are commonly used to determine the prevalence of Toxoplasma gondii infection in livestock, but the predictive value of seropositivity with respect to the presence of infective tissue cysts is less clear. The present study aimed at the identification of seropositive and seronegative free-range laying hens from organic and backyard farms, and the relationship with the presence of viable tissue cysts. In addition, potential risk and protective factors on the selected farms were investigated. An in-house T. gondii surface antigen (TgSAG1, p30, SRS29B) ELISA was validated with sera from experimentally infected chickens and used to examine 470 serum samples collected from laying hens from large organic and small backyard farms at the end of their laying period. A total of 11.7% (55/470) of all chickens tested positive, and another 18.9% (89/470) of test results were inconclusive. The highest seroprevalences were observed on small backyard farms with 47.7% (41/86) of chickens being seropositive while another 20.9% (18/86) of test results were inconclusive. Twenty-nine seropositive, 20 seronegative and 12 laying hens which yielded inconclusive ELISA results, were selected for further examination. Hearts and limb muscles of these hens were examined for T. gondii tissue cysts in a bioassay with IFNÉ£-knockout or IFNÉ£-receptor-knockout mice. Viable T. gondii was isolated from 75.9% (22/29) of the seropositive, 25.0% (3/12) of the inconclusive, and 5.0% (1/20) of the seronegative chickens. All 26 chickens tested positive in heart samples, while drumstick muscles (i.e. limb muscles) tested positive only in three. Data on putative risk and protective factors were collected on the farms using a standard questionnaire. Generalised multilevel modelling revealed farm size, cat related factors ('cats on the premise', 'cats used for rodent control'), hen house/hall related factors ('size category of hen house/hall', 'frequency category of cleaning hen house/hall', 'service period') as significantly associated with seropositivity to T. gondii in hens. The final model, which included the age of the birds as an effect modifier and farm as a random effect variable, revealed that the use of cats for rodent control and an area available per hen in the chicken run of ≥10sqm were statistically significant risk factors for T. gondii seropositivity. Overall this study showed that exposure to T. gondii is common in small backyard farms but is rare on large organic farms with a high density of free-range hens, even when cats were present on the premises.

[Visual associative memory and the orientation-contingent color after-effect]. / Zritel'naia assotsiativnaia pamiat' i éffekt orientatsionno-obuslovlennogo tsvetovogo posledeistviia.

The traditional explanation of the McCollough effect (ME) by selective adaptation of single detectors selective to color and orientation suffers from a number of inconsistencies: 1) the ME lasts much longer (from several days up to 3 months) than the ordinary adaptation, the decay of the effect being completely arrested by night sleep or occluding the eye for a long time; 2) the strength of the ME practically does not depend on the intensity of adapting light; and 3) a set of related pattern-contingent after-effects discovered later required for such an explanation new detectors, specific for other patterns. These properties can be explained, however, in the framework of associative memory and novelty filters. A computational model has been developed, which consists of 1) an input layer of two (left and right eyes) square matrices with two analog receptors (red and green) in each pixel, 2) an isomorphic associative neural layer, each analog neuron being synaptically connected with all receptors of both eyes, and 3) an output layer (novelty filter). The modification of synaptic efficacies conforms to the Hebb learning rule. The function of the model was examined by simulation. After a few presentations of colored gratings, the model displays the ME that is slowly destroyed by subsequent presentations of random pictures. With a sufficiently large receptor matrix, the effect lasts a thousand times longer than the period of adaptation. Continuous darkness does not change the strength of the effect. Like in real ME, the model does not display interocular transfer. The model can account for different pattern-contingent color after-effects without assuming any predetermined specific detectors. Such detectors are constructed in the course of adaptation to specific stimuli (gratings).

[Protection from virulent Salmonella groups B and D after the peroral immunization of chicks with a hybrid of Salmonella typhimurium and Salmonella dublin]. / Zashchita ot virulentnykh sal'monell grupp B i D posle peroral'noi immunizatsii tsypliat gibridom Salmonella typhimurium i Salmonella dublin.

Chickens over 10 days old, infected orally with virulent salmonellae, were found to remain alive. Histologic investigation showed the development of mild enteritis and more pronounced, lasting for more than two weeks, inflammation of the cecum, dissemination and focal lesions in the liver (granulomas, necrosis). In experiments on the oral immunization of 3-day old chickens the bivalent hybrid of S. typhimurium vaccine strain 274 and S. dublin induced only pronounced blast transformation in lymphatic follicles of the cecum, hyperplasia of activated macrophages and formation of granulomas from these macrophages and lymphocytes. After oral challenge of the immunized chickens with virulent salmonellae of group B (S. typhimurium) and group D (S. enteritidis, S. gallinarum-pullorum) the chickens exhibited sharply pronounced protection against adhesion, colonization and invasion, and a few penetrating bacteria were rapidly destroyed by immune macrophages. Hybrid strain 274/O9 proved to be suitable for use as oral bivalent vaccine against salmonellosis in chickens.

[The use of a live Salmonella bivalent vaccine for decreasing the circulation of salmonellae in a poultry plant]. / Primenenie zhivoi sal'monelleznoi bivalentnoi vaktsiny dlia snizheniia tsirkuliatsii sal'monell na ptitsefabrike.

S. typhimurium (expressing antigen 09 of S.dublin) recombinant vaccine strain was tested at a large poultry-breeding farm under the conditions of permanent circulation of salmonellae. The oral immunization of laying hens in three administrations with doses of 5 x 10(8), 5 x 10(9) and 5 x 10(9) microbial cells induced an increase in titers of antibodies only to group D salmonellae, while in immunized chickens (each receiving a dose of 3 x 10(8) microbial cells) antibodies to both group B and group D salmonellae were detected. The vaccination of laying hens and chickens produced a protective effect due to the development of immunity and the decrease of Salmonella contamination of eggs laid by immunized hens, thus reducing the risk of infection at incubator stations. Total mortality during the first 3 months of life was essentially lower (p < 0.001) among immunized chickens (4.41 +/- 0.09%) than in the control group of nonimmunized chickens (5.91 +/- 0.10%).

[Comparative effectiveness of various methods of fluid therapy in the treatment of moderate and severe burn shock]. / Sravnitel'naia éffektivnost' razlichnykh zhidkostnykh rezhimov pri lechenii ozhogovogo shoka legkoi i srednei stepeni.

Clinical study of the efficacy of four types of fluid treatment in mild and moderately severe burn shock in 134 patients showed that the prescription of oral alkaline solutions is sufficient in mild burn shock whereas in moderately severe shock fluids must be given by infusion. Intravenous infusion of glucose-electrolyte solutions is sufficiently effective in such cases.

[Treatment of burn shock with crystalloid solutions]. / Lechenie ozhogovogo shoka kristalloidnymi rastvorami.

The authors describe their experience of the treatment of burn shock in 124 patients with the square surface of the thermal injury about 20-49% of the body surface with the help of crystalloid solutions. The investigation of a number of parameters of hemodynamics, hemoconcentration, metabolism showed that patients with the Frank index 20-60 could be helped out of burn shock not only without blood and its albuminous preparations but also without infusions of dextran drugs. Transfusions of blood and its albuminous preparations are shown to be necessary in the nearest postshock period in patients with the Frank index 30-60. It is emphasized that the volume of intravenous infusions should be reduced in cases of associated lesions of the skin and the respiratory tracts.

Novel tools for the diagnosis and differentiation of acute and chronic bovine besnoitiosis.

Diagnosis of acute bovine besnoitiosis is a major diagnostic problem. We developed diagnostic tests to serologically diagnose and differentiate acute and chronic cases of bovine besnoitiosis using affinity purified antigens of Besnoitia besnoiti tachyzoites in immunoblots and in both, a conventional ELISA and an avidity ELISA. Sera of acutely and chronically infected cattle were investigated using these tests. Acutely infected cattle initially recognised an antigen of 74 kDa relative molecular mass, followed by reactions with increasing intensity against 81 and 28 kDa antigens. In addition, faint reactions against antigens with 36, 37, 39 and 42 kDa molecular mass started soon after seroconversion and increased over time. An antigen of 45 kDa molecular mass was transiently recognised early after infection but not or only weakly in the chronic stage. At least two antigens, the 39 and the 42 kDa antigens, seem to be located on the surface of B. besnoiti tachyzoites as determined by biotinylation. Affinity purified antigen was used to establish an APure-BbELISA which showed excellent sensitivity (100%) relative to a serological reference system in naturally, most likely chronically, infected cattle. Specificity was also high (99.8%) as determined in cattle from herds with Neospora caninum-associated abortions. The antibody levels in APure-BbELISA were correlated with the parasite load in the skin or the mucous membrane of the vestibulum vaginae as determined by real-time PCR. In acute cases of bovine besnoitiosis (confirmed by the detection of low avidity IgG in the APure-BbELISA) first specific antibodies were detected by ELISA in all animals except one, at the same time or earlier than in the serological reference system. The detection of parasite DNA in skin by real-time PCR was clearly superior to serological analysis in detecting infected cattle during acute besnoitiosis.