Isochronous island bifurcations driven by resonant magnetic perturbations in tokamaks.

Recent evidence shows that heteroclinic bifurcations in magnetic islands may be caused by the amplitude variation of resonant magnetic perturbations in tokamaks. To investigate the onset of these bifurcations, we consider a large aspect ratio tokamak with an ergodic limiter composed of two pairs of rings that create external primary perturbations with two sets of wave numbers. An individual pair produces hyperbolic and elliptic periodic points, and its associated islands, that are consistent with the Poincaré-Birkhoff fixed-point theorem. However, for two pairs producing external perturbations resonant on the same rational surface, we show that different configurations of isochronous island chains may appear on phase space according to the amplitude of the electric currents in each pair of the ergodic limiter. When one of the electric currents increases, isochronous bifurcations take place and new islands are created with the same winding number as the preceding islands. We present examples of bifurcation sequences displaying (a) direct transitions from the island chain configuration generated by one of the pairs to the configuration produced by the other pair, and (b) transitions with intermediate configurations produced by the limiter pairs coupling. Furthermore, we identify shearless bifurcations inside some isochronous islands, originating nonmonotonic local winding number profiles with associated shearless invariant curves.

Three Months of Composting Are Enough to Eliminate the Fowl Typhoid Bacteria

The composting technique has been increasingly highlighted in poultry production units, as an efficient and low-cost solution for the destination of carcasses. The process is based on the accelerated decomposition of organic material under high temperatures, associated with eliminating pathogenic microorganisms. This study aims to evaluate the effectiveness and the time necessary for the elimination of Salmonella Gallinarum in carcasses of poultry submitted to the composting process. The composting was carried out following the models used in the field, and microbiological analysis was performed in five different periods: 45, 90, 120, 150 and 180-days after closing the composter. After 90 days of experiment and in the subsequent analysis, the elimination of the bacteria in 100% of the samples was verified, validating the composting process as an effective method for eliminating S. Gallinarum in poultry carcasses, when respecting the period necessary for the elimination of the bacteria and the good quality of the structure adopted for the process.(AU)

Inclusion of Organic Acids in the Drinking Water and Feed for the Control of Salmonella Heidelberg in Broilers

Poultry products may be a source of foodborne human salmonellosis. The use of alternatives to antimicrobials that are not harmful to humans may reduce the presence of Salmonella spp. in poultry production. Among the products used, organic acids stand out. In the present study, three different organic acid (OA) blends were evaluated for the control of Salmonella Heidelberg (SH) in commercial broilers. Day-old chicks (n = 114) were randomly assigned to four treatments, with three replicates of 12 birds each. Birds in treatments A and B received SCFA (0.2mL/L) and SCFA + MCFA (0.2mL/L), respectively, in the drinking water, while birds in treatment C received SCFA + MCFA in the feed (2g/Kg of feed). Birds from treatment D did not receive OAs (control group). At 8 days of age, each bird was orally inoculated with SH at 108 CFU/mL, and cloacal swabs and SH enumeration of the cecal content were performed 24-, 48-, and 72-hours post-inoculation (hpi). The results show a reduction of both SH shedding and counts in the birds fed OAs at all pi times relative to the control birds. Fecal shedding was significantly lower in the OA-treated groups compared with the control group. As for SH presence in the cecum, significant differences were detected between groups C and D at 24 and 72 hpi, and between groups B and D at 72 hpi. The results of this study indicate that the use of feeding OAs to broilers may contribute to reduce the incidence of SH in the poultry production chain, allowing better flock health management, provided an efficient biosecurity program is employed.(AU)

Deciphering the role of ttrA and pduA genes for Salmonella enterica serovars in a chicken infection model.

Salmonella enterica serovars use self-induced intestinal inflammation to increase electron acceptor availability and to obtain a growth advantage in the host gut. There is evidence suggesting that the ability of Salmonella to use tetrathionate and 1,2-propanediol provides an advantage in murine infection. Thus, we present here the first study to evaluate both systemic infection and faecal excretion in commercial poultry challenged by Salmonella Enteritidis (SE) and S. Typhimurium (STM) harbouring deletions in ttrA and pduA genes, which are crucial to the metabolism of tetrathionate and 1,2-propanediol, respectively. Mutant strains were excreted at higher rates when compared to the wild-type strains. The highest rates were observed with white egg-layer and brown egg-layer chicks (67.5%), and broiler chicks (56.7%) challenged by SEΔttrAΔpduA, and brown egg-layer chicks (64.8%) challenged by STMΔttrAΔpduA. SEΔttrAΔpduA presented higher bacterial counts in the liver and spleen of the three chicken lineages and caecal contents from the broiler chickens, whereas STMΔttrAΔpduA presented higher counts in the liver and spleen of the broiler and brown-egg chickens for 28 days post-infection (P < 0.05). The ttrA and pduA genes do not appear to be major virulence determinants in faecal excretion or invasiveness for SE and STM in chickens. RESEARCH HIGHLIGHTSttrA and pudA do not impair gut colonization or systemic infection in chicks.Mutant strains were present in higher numbers in broilers than in laying chicks.Mutants of SE and STM showed greater pathogenicity in broiler chicks than layers.

Evaluation of propanediol and cobalamin metabolism in the intestinal colonization and systemic invasion of Salmonella Enteritidis in laying hens / [Avaliação do metabolismo do propanodiol e da cobalamina na colonização intestinal e na infecção sistêmica de Salmonella Enteritidis em poedeiras]

Embora Salmonella Enteritidis (SE) seja capaz de metabolizar 1,2-propanodiol (1,2-Pd), utilizado como fonte de carbono e de energia ao longo de uma rota dependente de vitamina B12, a importância deste composto na infeção de Gallus gallus domesticus por SE permanece desconhecida. No presente estudo, foram construídos um mutante de SE sem os genes pduCDE, que codifica a propanodiol desidratase (Pdu), e outro contendo as deleções no pduCDE e também nos genes cobS e cbiA, responsáveis pela síntese de vitamina B12. Em seguida, avaliou-se a importância do metabolismo do 1,2-Pd em SE para colonização intestinal de infecção sistêmica de poedeiras comerciais. As estirpes mutantes de SE foram capazes de colonizar o intestino, de serem excretadas nas fezes e de invadir o baço e o fígado na mesma intensidade que a estirpe selvagem, o que sugere que os produtos dos genes pduC, pduD, pduE, cobS e cbiA não são essenciais durante infecção por Salmonella Enteritidis nessa espécie.(AU)

Antimicrobial Resistance in Salmonella Serovars Isolated From an Egg-Producing Region in Brazil

Fowl paratyphoid infections are caused by different Salmonella serovars that can affect a wide range of hosts. Due to its complex epidemiology, Salmonella serovar identification is crucial for the development and implementation of monitoring and control programs in poultry farms. Moreover, the characterization of the antimicrobial resistance profiles of Salmonella strains isolated from livestock is relevant to public health because they are a common causative agent of foodborne diseases. The objective of this study was to investigate the presence of Salmonella spp. and to identify the antimicrobial resistance profiles of strains isolated in the midwestern region of São Paulo state, which accounts for the highest production of table eggs in Brazil. For this purpose, 2008 fecal samples were collected on 151 commercial layer farms and submitted to microbiological analyses. Twenty-two serovars were isolated from 80 (52.9%) farms, among which S. Mbandaka and S. Braenderup were the most prevalent. All isolates expressed resistance to at least one of the 23 antimicrobials tested, and the highest resistance rates were determined against streptomycin (93.5%) and sulfonamide (84.6%). Moreover, multidrug resistance was observed in 41% of the isolates and the maximum drug resistance profile was against ten different antimicrobials. Therefore, the identification of Salmonella serovars in poultry production provides epidemiological knowledge to develop prevention and control measures in order to ensure poultry health and to prevent human infection by multiresistant strains.

Antimicrobial Resistance in Salmonella Serovars Isolated From an Egg-Producing Region in Brazil

Fowl paratyphoid infections are caused by different Salmonella serovars that can affect a wide range of hosts. Due to its complex epidemiology, Salmonella serovar identification is crucial for the development and implementation of monitoring and control programs in poultry farms. Moreover, the characterization of the antimicrobial resistance profiles of Salmonella strains isolated from livestock is relevant to public health because they are a common causative agent of foodborne diseases. The objective of this study was to investigate the presence of Salmonella spp. and to identify the antimicrobial resistance profiles of strains isolated in the midwestern region of São Paulo state, which accounts for the highest production of table eggs in Brazil. For this purpose, 2008 fecal samples were collected on 151 commercial layer farms and submitted to microbiological analyses. Twenty-two serovars were isolated from 80 (52.9%) farms, among which S. Mbandaka and S. Braenderup were the most prevalent. All isolates expressed resistance to at least one of the 23 antimicrobials tested, and the highest resistance rates were determined against streptomycin (93.5%) and sulfonamide (84.6%). Moreover, multidrug resistance was observed in 41% of the isolates and the maximum drug resistance profile was against ten different antimicrobials. Therefore, the identification of Salmonella serovars in poultry production provides epidemiological knowledge to develop prevention and control measures in order to ensure poultry health and to prevent human infection by multiresistant strains.(AU)

Duplex Real-Time PCR Using Sybr Green I for Quantification and Differential Diagnosis between Salmonella Enteritidis and Salmonella Typhimurium

The incidence of foodborne diseases caused by the genus Salmonella spp. in industrialized countries is often high in epidemiological surveys. Obtaining a rapid diagnostic test for identification of bacteria is crucial in order to rapidly implement control measures to contain bacterial spread, to reduce losses in animal production and to avoid risks from food-borne infections to human health. The aim of this study was to standardize duplex real-time PCR using SYBr Green I for differential and quantitative diagnosis of S. Typhimurium and S. Enteritidis. According to the experiment, the melting temperature of 85°C was observed for a 206bp amplified product when S. Enteritidis DNA was added to the reaction. S. Typhimurium DNA showed that the melting temperature of 79°C when observed for a 62bp amplified product. The standard curve showed the high sensitivity of the proposed test, since it was possible to obtain eight quantification points, starting at 108 CFU/mL and ending at 101 CFU/mL. As a result of the present study, a real-time PCR duplex reaction with high sensitivity, specificity and based on the fluorescence of SYBr Green I was standardized. In addition, this methodology aligns low cost to the faster diagnostic result, in relation to other molecular tests, making it attractive for application in routine laboratory analyzes.

Duplex Real-Time PCR Using Sybr Green I for Quantification and Differential Diagnosis between Salmonella Enteritidis and Salmonella Typhimurium

The incidence of foodborne diseases caused by the genus Salmonella spp. in industrialized countries is often high in epidemiological surveys. Obtaining a rapid diagnostic test for identification of bacteria is crucial in order to rapidly implement control measures to contain bacterial spread, to reduce losses in animal production and to avoid risks from food-borne infections to human health. The aim of this study was to standardize duplex real-time PCR using SYBr Green I for differential and quantitative diagnosis of S. Typhimurium and S. Enteritidis. According to the experiment, the melting temperature of 85°C was observed for a 206bp amplified product when S. Enteritidis DNA was added to the reaction. S. Typhimurium DNA showed that the melting temperature of 79°C when observed for a 62bp amplified product. The standard curve showed the high sensitivity of the proposed test, since it was possible to obtain eight quantification points, starting at 108 CFU/mL and ending at 101 CFU/mL. As a result of the present study, a real-time PCR duplex reaction with high sensitivity, specificity and based on the fluorescence of SYBr Green I was standardized. In addition, this methodology aligns low cost to the faster diagnostic result, in relation to other molecular tests, making it attractive for application in routine laboratory analyzes.(AU)

Phylogeny and variability of Colletotrichum truncatum associated with soybean anthracnose in Brazil.

AIMS: Fungal diseases are among the main factors limiting high yields of soybean crop. Colletotrichum isolates from soybean plants with anthracnose symptoms were studied from different regions and time periods in Brazil using molecular, morphological and pathogenic analyses. METHODS AND RESULTS: Bayesian phylogenetic inference of GAPDH, HIS3 and ITS-5.8S rDNA sequences, the morphologies of colony and conidia, and inoculation tests on seeds and seedlings were performed. All isolates clustered only with Colletotrichum truncatum species in three well-separated clusters. Intraspecific genetic diversity revealed 27 distinct haplotypes in 51 fungal isolates; some of which were identical to C. truncatum sequences from other regions around the world, while others were related to alternative hosts. Conidia were falcate, hyaline, unicellular and aseptate, formed in acervuli, with variable dimensions. Despite being pathogenic to seedlings by both inoculation methods, variation was observed in the aggressiveness of the tested isolates, which was not correlated with genetic variation. CONCLUSION: The identification of C. truncatum in the sampled isolates was evidenced as being the only causal agent of soybean anthracnose in Brazil until 2007, with relevant genetic, morphological and pathogenic variability as well as a broad geographical origin. The wide distribution of the predominant C. truncatum haplotype indicated the existence of a highly efficient mechanism of pathogen dispersal over long distances, reinforcing the role of seeds as the primary source of disease inoculum. SIGNIFICANCE AND IMPACT OF THE STUDY: The characterization and distribution of Colletotrichum species in soybean-producing regions in Brazil is fundamental for understanding the disease epidemiology and for ensuring effective control strategies against anthracnose.

Experimental infection of chickens by a flagellated motile strain of Salmonella enterica serovar Gallinarum biovar Gallinarum.

Salmonella enterica subsp. enterica serovar Gallinarum biovar Gallinarum (SG) causes fowl typhoid (FT), a septicaemic disease which can result in high mortality in poultry flocks. The absence of flagella in SG is thought to favour systemic invasion, since bacterial recognition via Toll-like receptor (TLR)-5 does not take place during the early stages of FT. In the present study, chicks susceptible to FT were inoculated with a wild type SG (SG) or its flagellated motile derivative (SG Fla(+)). In experiment 1, mortality and clinical signs were assessed, whereas in experiment 2, gross pathology, histopathology, systemic invasion and immune responses were evaluated. SG Fla(+) infection resulted in later development of clinical signs, lower mortality, lower bacterial numbers in the liver and spleen, and less severe pathological changes compared to SG. The CD8(+) T lymphocyte population was higher in the livers of chicks infected with SG at 4 days post-inoculation (dpi). Chicks infected with SG had increased expression of interleukin (IL)-6 mRNA in the caecal tonsil at 1 dpi and increased expression of IL-18 mRNA in the spleen at 4 dpi. In contrast, the CD4(+) T lymphocyte population was higher at 6 dpi in the livers of birds infected with SG Fla(+). Therefore, flagella appeared to modulate the chicken immune response towards a CD4(+) T profile, resulting in more efficient bacterial clearance from systemic sites and milder infection.

Complete genome sequence of bean rugose mosaic virus, genus Comovirus.

Since the first report in Costa Rica in 1971, bean rugose mosaic virus (BRMV) has been found in Colombia, El Salvador, Guatemala and Brazil. In this study, the complete genome sequence of a soybean isolate of BRMV from Paraná State, Brazil, was determined. The BRMV genome consists of two polyadenylated RNAs. RNA1 is 5909 nucleotides long and encodes a single polypeptide of 1856 amino acids (aa), with an estimated molecular weight of 210 kDa. The RNA1 polyprotein contains the polypeptides for viral replication and proteolytic processing. RNA2 is 3644 nucleotides long and codes for a single polypeptide of 1097 aa, containing the movement and coat proteins. This is the first complete genome sequence of BRMV. When compared with available aa sequences of comoviruses, the highest identities of BRMV coat proteins and proteinase polymerase were 57.5 and 58 %, respectively. These were below the 75 and 80 % identity limits, respectively, established for species demarcation in the genus. This confirms that BRMV is a member of a distinct species in the genus Comovirus.

Viability of Yersinia pestis subcultures in agar stabs.

UNLABELLED: Since its identification as the causative agent of plague in 1894, thousands of Yersinia pestis strains have been isolated and stored. Here, we report the ability of Y. pestis to survive up to 47 years in agar stabs, in rubber-stoppered tubes, under refrigeration (+4 to +10°C), although overall subculture recovery rates were poor and inversely related to the length of time stored. Genetic characterization of virulence gene presence among these subcultures was suggestive of significant variation in the genomic stability of Y. pestis subcultures stored under these conditions. Specifically, we found variation in the presence of plasmid and chromosomal virulence markers (genes pla, lcrV, caf1 and irp2) among multiple subcultures of Y. pestis strains in the 'Collection of Yersinia pestis' (Fiocruz-CYP) maintained by the SRP of FIOCRUZ-PE in Brazil. This variation, together with all of the inherent temporal, geographic and other genetic variation represented by all of the recoverable strains in this historical collection was preserved in new frozen culture stocks stored at -70°C as a result of this study. These frozen culture stocks represent a valuable resource for future comparative studies of Y. pestis. SIGNIFICANCE AND IMPACT OF THE STUDY: We report the ability of Yersinia pestis to survive up to 47 years in agar stabs, in rubber-stoppered tubes, under refrigeration (+4 to +10°C), although overall subculture recovery rates were poor and inversely related to the length of time stored. Genetic characterization of virulence gene presence among these subcultures was suggestive of significant variation in the genomic stability of Y. pestis subcultures stored under these conditions. This variation, together with all of the inherent temporal, geographic and other genetic variation represented by all of the recoverable strains in the historical 'Collection of Yersinia pestis' (Fiocruz-CYP) maintained by the SRP of FIOCRUZ-PE in Brazil was preserved in new frozen culture stocks stored at -70°C as a result of this study. These frozen culture stocks represent a valuable resource for future comparative studies of Y. pestis.

Evaluation of Disinfectants Used in Pre-Chilling water Tanks of Poultry Processing Plants

In poultry processing plants, disinfectants are often added to pre-chilling water tanks to reduce microbial contamination. The present study aimed at evaluating the effect of five disinfectants (acidified sodium chlorite, alkyl dimethyl benzyl ammonium chloride, chlorine dioxide, peracetic acid, and sodium hypochlorite) on the populations of food quality indicator microorganisms and on Salmonella Enteritidis (SE) in the presence and absence of organic matter. The results showed that chlorine dioxide and sodium hypochlorite did not reduce microbial carcass counts. On the other hand, acidified sodium chlorite, alkyl dimethyl benzyl ammonium chloride and peracetic acid reduced total and fecal coliform counts. Peracetic acid reduced the number of psychrotrophic microorganisms. All products were effective in reducing SE counts only in the absence of organic matter. Acidified sodium chlorite, alkyl dimethyl benzyl ammonium chloride and peracetic acid could be candidates for the replacement of sodium hypochlorite (commonly used in Brazil) in pre-chilling tanks. (AU)

Evaluation of Disinfectants Used in Pre-Chilling water Tanks of Poultry Processing Plants

In poultry processing plants, disinfectants are often added to pre-chilling water tanks to reduce microbial contamination. The present study aimed at evaluating the effect of five disinfectants (acidified sodium chlorite, alkyl dimethyl benzyl ammonium chloride, chlorine dioxide, peracetic acid, and sodium hypochlorite) on the populations of food quality indicator microorganisms and on Salmonella Enteritidis (SE) in the presence and absence of organic matter. The results showed that chlorine dioxide and sodium hypochlorite did not reduce microbial carcass counts. On the other hand, acidified sodium chlorite, alkyl dimethyl benzyl ammonium chloride and peracetic acid reduced total and fecal coliform counts. Peracetic acid reduced the number of psychrotrophic microorganisms. All products were effective in reducing SE counts only in the absence of organic matter. Acidified sodium chlorite, alkyl dimethyl benzyl ammonium chloride and peracetic acid could be candidates for the replacement of sodium hypochlorite (commonly used in Brazil) in pre-chilling tanks.

Animal board invited review: advances in proteomics for animal and food sciences.

Animal production and health (APH) is an important sector in the world economy, representing a large proportion of the budget of all member states in the European Union and in other continents. APH is a highly competitive sector with a strong emphasis on innovation and, albeit with country to country variations, on scientific research. Proteomics (the study of all proteins present in a given tissue or fluid - i.e. the proteome) has an enormous potential when applied to APH. Nevertheless, for a variety of reasons and in contrast to disciplines such as plant sciences or human biomedicine, such potential is only now being tapped. To counter such limited usage, 6 years ago we created a consortium dedicated to the applications of Proteomics to APH, specifically in the form of a Cooperation in Science and Technology (COST) Action, termed FA1002--Proteomics in Farm Animals: www.cost-faproteomics.org. In 4 years, the consortium quickly enlarged to a total of 31 countries in Europe, as well as Israel, Argentina, Australia and New Zealand. This article has a triple purpose. First, we aim to provide clear examples on the applications and benefits of the use of proteomics in all aspects related to APH. Second, we provide insights and possibilities on the new trends and objectives for APH proteomics applications and technologies for the years to come. Finally, we provide an overview and balance of the major activities and accomplishments of the COST Action on Farm Animal Proteomics. These include activities such as the organization of seminars, workshops and major scientific conferences, organization of summer schools, financing Short-Term Scientific Missions (STSMs) and the generation of scientific literature. Overall, the Action has attained all of the proposed objectives and has made considerable difference by putting proteomics on the global map for animal and veterinary researchers in general and by contributing significantly to reduce the East-West and North-South gaps existing in the European farm animal research. Future activities of significance in the field of scientific research, involving members of the action, as well as others, will likely be established in the future.

Phylogeny and micro-habitats utilized by lizards determine the composition of their endoparasites in the semiarid Caatinga of Northeast Brazil.

Trophic networks can have architectonic configurations influenced by historical and ecological factors. The objective of this study was to analyze the architecture of networks between lizards, their endoparasites, diet, and micro-habitat, aiming to understand which factors exert an influence on the composition of the species of parasites. All networks showed a compartmentalized pattern. There was a positive relation between diet and the diversity of endoparasites. Our analyses also demonstrated that phylogeny and the use of micro-habitat influenced the composition of species of endoparasites and diet pattern of lizards. The principal factor that explained the modularity of the network was the foraging strategy, with segregation between the "active foragers" and "sit-and-wait" lizards. Our analyses also demonstrated that historical (phylogeny) and ecological factors (use of micro-habitat by the lizards) influenced the composition of parasite communities. These results corroborate other studies with ectoparasites, which indicate phylogeny and micro-habitat as determinants in the composition of parasitic fauna. The influence of phylogeny can be the result of coevolution between parasites and lizards in the Caatinga, and the influence of micro-habitat should be a result of adaptations of species of parasites to occupy the same categories of micro-habitats as hosts, thus favoring contagion.

Salmonella serovars in laying hen flocks and commercial table eggs from a region of São Paulo state, Brazil

Salmonella spp. is the main originator of human foodborne diseases worldwide and is mainly transmitted by food containing eggs. In Brazil, as a result of the lack of studies and data collection very little is known about the prevalence of Salmonella spp. in laying hen flocks and commercial table eggs. Consequently the present study was elaborated and aimed at generating data about Salmonella spp. in part of the Brazilian egg production chain. Eight flocks of day-old chicks, eight flocks of adult laying hens (four vaccinated with bacterin against Salmonella Enteritidis and four unvaccinated) and commercial table eggs from four supermarkets were examined. Salmonella spp. was isolated in 50 % of the newly hatched chicks, 25 % of the adult flocks and 1.5 % of egg samples examined. S. enterica subsp. enterica 4,12:r:-, S. Mbandaka, S. enterica subsp. enterica 6,7: z10:-, S. Enteritidis and S. Havana were the serovars isolated in birds. In commercial table-eggs S. Mbandaka, S. enterica subsp. enterica 6,7: z10:- and S. Braenderup were isolated. These results show that Salmonella spp. is present in laying hen flocks and consequently in eggs destined for human consumption. Probably, some of the Salmonella serovars are being introduced in egg farms by vertical via.

Salmonella serovars in laying hen flocks and commercial table eggs from a region of São Paulo state, Brazil

Salmonella spp. is the main originator of human foodborne diseases worldwide and is mainly transmitted by food containing eggs. In Brazil, as a result of the lack of studies and data collection very little is known about the prevalence of Salmonella spp. in laying hen flocks and commercial table eggs. Consequently the present study was elaborated and aimed at generating data about Salmonella spp. in part of the Brazilian egg production chain. Eight flocks of day-old chicks, eight flocks of adult laying hens (four vaccinated with bacterin against Salmonella Enteritidis and four unvaccinated) and commercial table eggs from four supermarkets were examined. Salmonella spp. was isolated in 50 % of the newly hatched chicks, 25 % of the adult flocks and 1.5 % of egg samples examined. S. enterica subsp. enterica 4,12:r:-, S. Mbandaka, S. enterica subsp. enterica 6,7: z10:-, S. Enteritidis and S. Havana were the serovars isolated in birds. In commercial table-eggs S. Mbandaka, S. enterica subsp. enterica 6,7: z10:- and S. Braenderup were isolated. These results show that Salmonella spp. is present in laying hen flocks and consequently in eggs destined for human consumption. Probably, some of the Salmonella serovars are being introduced in egg farms by vertical via.(AU)