Detection of Salmonella spp. in wild and domestic birds in an anthropized ecotone between the Cerrado and the Amazon Forest in Brazil.

Emergence of zoonotic infectious diseases represent one of the main threats to people worldwide. To properly understand and prevent zoonoses is fundamental to study their epidemiology and the possibility of spillover events, especially for commercially intensive domestic animals and humans. Here, we studied 210 wild birds from the "Ipucas" region, which consists of fragments of the Amazon Forest interspersed with fragments of the "Cerrado" that is subject to seasonal flooding and 75 domestic birds from neighboring poultry farming. Then, we molecularly diagnosed Salmonella and Chlamydia from wild birds and poultry. Among the wild birds, four were diagnosed with Chlamydia psittaci and 23 with Salmonella spp., while we detected 15 poultry infected by Salmonella spp. and no poultry with C. psittaci. We highlighted the common infections of wild and domestic birds in an anthropologically modified environment and potential spillover of Salmonella pathogens among wild and livestock birds. Those infections can harm the health of native and domestic species.

Stainless steel weld metal enhanced with carbon nanotubes.

This paper aims to establish the most indicated route to manufacture a nanostructured powder composed of 5 wt% Multi-walled Carbon Nanotubes and 304LSS powder. Four specimens were prepared using Mechanical Alloying and Chemical Treatment (CT) with Hydrogen Peroxide ([Formula: see text]) as the main processes. A thermal treatment post-processing was used in half of the samples to remove the remaining amorphous carbon and to evaluate its effects. Regarding the powder analysis, attachment, amorphous carbon degree, crystallinity, and doping of the CNT throughout the metal matrix were investigated. The nanostructured powders were then inserted as a core in a 304LSS tubular rod to perform the arc welding process. The CT route eliminated the amorphous carbon and generated more refiner grains, which provided a cross-section hardness gain of more than 40% regarding the 304LSS joint. In summary, the CT route, combined with the GTAW process, provided a new method for nanocomposite manufacturing by combining shorter preparation steps, obtaining an improvement in the microstructural and hardness performance.

High fidelity defines the temporal consistency of host-parasite interactions in a tropical coastal ecosystem.

Host-parasite interactions represent a selective force that may reduce hosts' lifespan, their reproductive success and survival. Environmental conditions can affect host-parasite communities, leading to distinct patterns of interactions with divergent ecological and evolutionary consequences for their persistence. Here, we tested whether climatic oscillation shapes the temporal dynamics of bird-haemosporidian associations, assessing the main mechanisms involved in the temporal dissimilarity of their interactions' networks. For two years, we monthly sampled birds in a tropical coastal ecosystem to avian malaria molecular diagnosis. The studied networks exhibited high specialization, medium modularity, with low niche overlap among parasites lineages. Moreover, alpha and ß-diversity of hosts, parasites and their interactions, as well as the structure of their networks were temporally consistent, i.e., stable under fluctuations in temperature or precipitation over seasons. The structure and temporal consistency of the studied antagonistic networks suggest a high fidelity between partners, which is likely relevant for their evolutionary persistence.

Malaria in penguins - current perceptions.

Avian malaria is a mosquito-borne disease caused by protozoans of the genus Plasmodium, and it is considered one of the most important causes of morbidity and mortality in captive penguins, both in zoological gardens and rehabilitation centres. Penguins are known to be highly susceptible to this disease, and outbreaks have been associated with mortality as high as 50-80% of affected captive populations within a few weeks. The disease has also been reported in wild penguin populations, however, its impacts on the health and fitness of penguins in the wild is not clear. This review provides an overview of the aetiology, life cycle and epidemiology of avian malaria, and provides details on the strategies that can be employed for the diagnostic, treatment and prevention of this disease in captive penguins, discussing possible directions for future research.

Prevalence and diversity of avian malaria parasites in migratory Black Skimmers (Rynchops niger, Laridae, Charadriiformes) from the Brazilian Amazon Basin.

The Medium Solimões River region in the Brazilian Amazon Basin is an area utilized for reproduction and nesting by a variety of species of migratory aquatic birds such as Black Skimmers (Rynchops niger). These migratory birds form mixed-species reproductive colonies with high population densities and exhibit a large range of migration routes. This study aimed to describe the prevalence and diversity of the avian malaria parasites Plasmodium and Haemoproteus in Black Skimmers, on the basis of the association between microscopic observation of blood smears and amplification of the mitochondrial cytochrome b gene (mtDNA cyt-b). The overall prevalence rates of the parasites for juvenile and adult bird specimens were 16% (5/31) and 22% (15/68), respectively. Sequencing the mtDNA cyt-b marker revealed two Plasmodium lineages, which had been previously described in different regions of the American continent, including a Neotropical region in Southeast Brazil, and one Haemoproteus lineage. The fact that avian malarial parasites have been found infecting the Black Skimmers in the Brazilian Amazon ecosystem, which exhibits considerable diversity, highlights the importance of these migratory birds as a potential source of infection and dispersion of pathogens to other susceptible birds of the Nearctic and Neotropical regions.

Do ticks and Borrelia burgdorferi s.l. constitute a burden to birds?

Ticks consume resources from their hosts shaping their life-history traits and are vectors of many zoonotic pathogens. Several studies have focused on the health effects of blood-sucking ectoparasites on avian hosts, but there is limited information on the effects of ticks on adult and sub-adult birds, which may actively avoid ticks and are likely to present low infestation intensities. We evaluated the effects of the presence of feeding ticks and intensity of infestation on health variables of avian hosts. We also evaluated whether these variables were affected by tick infection by Borrelia burgdorferi sensu lato (s.l.) and by the presence of Borrelia infection on the birds' skin. Presence of parasite association among ticks, haemosporidea and Borrelia within the bird-host was also tested. We found that infestation by ticks significantly increased heterophyl/lymphocyte ratio in Turdus merula suggesting increased stress. This was especially evident at high infestation intensities when a significant decrease in body mass and body condition (body mass corrected for size) was also observed. Erithacus rubecula infested with more than 10 larvae tended to have lower haematocrit and blood haemoglobin. Plasma globulin concentration in T. merula tended to be affected by the presence of attached ticks and their infection with Borrelia, but this depended on the age of the bird. No association was detected among ticks, haemosporidea and Borrelia infection. We showed that ticks have detrimental effects on their avian hosts even under natural infestation conditions and that confirmed Borrelia reservoir hosts may also present symptoms of infection, though these may be subtle.

Seroprevalence of orthopoxvirus in an Amazonian rural village, Acre, Brazil.

Vaccinia virus strains from the family Poxviridae have been frequently isolated in Brazil and associated with outbreaks of exanthematic disease affecting cows and humans. An ELISA IgG was applied to evaluate the seroprevalence of orthopoxviruses in a community located in a rural settlement in the Amazon region, where no orthopoxvirus outbreaks have yet been reported. An overall seroprevalence of 27.89% was found, and it was 23.38% in the non-vaccinated population (smallpox vaccination). These results strongly suggest that orthopoxviruses circulate in this population, and it is the first finding of seropositivity for orthopoxviruses in a population without any previously reported outbreaks.

Naturally acquired inhibitory antibodies to Plasmodium vivax Duffy binding protein are short-lived and allele-specific following a single malaria infection.

The Duffy binding protein of Plasmodium vivax (DBP) is a critical adhesion ligand that participates in merozoite invasion of human Duffy-positive erythrocytes. A small outbreak of P. vivax malaria, in a village located in a non-malarious area of Brazil, offered us an opportunity to investigate the DBP immune responses among individuals who had their first and brief exposure to malaria. Thirty-three individuals participated in the five cross-sectional surveys, 15 with confirmed P. vivax infection while residing in the outbreak area (cases) and 18 who had not experienced malaria (non-cases). In the present study, we found that only 20% (three of 15) of the individuals who experienced their first P. vivax infection developed an antibody response to DBP; a secondary boosting can be achieved with a recurrent P. vivax infection. DNA sequences from primary/recurrent P. vivax samples identified a single dbp allele among the samples from the outbreak area. To investigate inhibitory antibodies to the ligand domain of the DBP (cysteine-rich region II, DBP(II)), we performed in vitro assays with mammalian cells expressing DBP(II) sequences which were homologous or not to those from the outbreak isolate. In non-immune individuals, the results of a 12-month follow-up period provided evidence that naturally acquired inhibitory antibodies to DBP(II) are short-lived and biased towards a specific allele.

Avian malaria in captive psittacine birds: detection by microscopy and 18S rRNA gene amplification.

A cross-sectional survey was conducted to estimate the occurrence of malaria infection among captive psittacine birds (n=127) from three zoological gardens in Brazil. Malaria infection was evaluated by the association of direct examination of blood smears with amplification of the 18SSU rRNA gene of the Plasmodium genus, demonstrating an overall occurrence of 36%. Most infected bird species were Amazona aestiva (28/73), Ara ararauna (6/10), and Amazona amazonica (3/10). The low parasitemias observed among the infected birds suggest a chronic infection. The sequence analyses of 10 isolates indicate a potential occurrence of four distinct Plasmodium lineages. These findings provide new data on malarial infection in captive psittacine birds, and emphasize the need for better control of importation and exportation of these birds.

Inhibitory properties of the antibody response to Plasmodium vivax Duffy binding protein in an area with unstable malaria transmission.

The function of the Plasmodium vivax Duffy binding protein (DBP) during the erythrocyte invasion process is critical for successful parasite growth and pathogenesis in human infections. Although DBP is the subject of intensive malaria vaccine research, investigations on the functional proprieties of anti-DBP antibodies in the human population have been limited [Infect Immun68 (2000) 3164]. In the present study, we examined the ability of sera from different populations of the Brazilian Amazon--an area of markedly unstable malaria transmission--to inhibit the erythrocyte-binding function of the DBP ligand domain (region II, DBP(II)). We found that long-term exposure to malaria in the Amazon area elicits DBP-specific antibodies that inhibit the binding of different DBP(II) variants to erythrocytes. Despite the great variability of inhibitory antibody responses observed among study participants, we observed a positive correlation between erythrocyte binding-inhibitory activity and enzyme-linked immunosorbent assay anti-DBP antibodies. Of importance, there was a non-significant tendency towards increased levels of anti-DBP antibodies among individuals with asymptomatic P. vivax infections.

Aspectos clínicos e imunopatológicos da ceratoplastia com membrana amniótica xenógena fresca e conservada em glicerina: estudo experimental em coelhos / Clinical and immunopatho logical aspects of keratoplasty with fresh and preserved in glycerin xenogenous amniotic membrane: experimental study in rabbits

Avaliaram-se, por meio da análise clínica, histopatologia e imunoistoquímica, os efeitos da aplicação da membrana amniótica xenógena fresca e conservada em glicerina, sobre os mecanismos imunológicos da superfície ocular. Para tal, utilizaram-se 40 coelhos, distribuídos em dois grupos experimentais, os quais foram avaliados por 21 dias. A avaliação clínica revelou que a membrana amniótica xenógena conservada em glicerina estimulou uma resposta inflamatória aguda maior que a membrana aplicada fresca. A análise histopatológica indicou que ambas se comportaram de forma semelhante a partir da primeira semana de pós-operatório, apresentando as alterações clássicas da resposta inflamatória da córnea, com o predomínio de infiltrado do tipo polimorfonuclear. A análise imunoistoquímica indicou que, ainda aos 21 dias, a resposta imune local é inespecífica, permitindo concluir que a resposta imune específica na córnea é tardia e que a córnea é um sítio privilegiado para aplicação de enxertos com características imunológicas diferentes, visto que não houve o estímulo para o desenvolvimento de uma resposta mais específica nos grupos avaliados durante toda a execução do experimento.

The dynamics of the inflammatory response and the mechanism involved in the healing process of cornea treated by keratoplasty, applying fresh and glycerin preserved xenogenous amniotic membranes as method to cover experimental ulcers were studied using 40 rabbits. The animals were allotted into two groups and evaluated during 21 days. The eyes were evaluated by histopathological study and immunohistochemical reaction. The clinical evaluation showed that the xenogenous amniotic membrane preserved in glycerin stimulated a greater acute inflammatory response than the one caused by fresh membrane. The histopathological analysis indicated that both membranes reacted in a very similar way from the first week post-surgery, presenting the classical alterations of the inflammatory response on the cornea. The immunohistochemical technique indicated that in no moment of the observation, the local immunologic response was specific. It was concluded that the specific immunologic response on the cornea took place later and that is a privileged site to use grafts with different immunological characteristics, once there was not a stimulus to the development of a more specific response in none of the evaluated groups throughout the experiment.

Avian malaria in Brazilian passerine birds: parasitism detected by nested PCR using DNA from stained blood smears.

The microscopical examination of Giemsa-stained thin blood smears and a nested PCR were performed to detect avian Plasmodium in 275 passerine birds from small and large fragments of Atlantic Forest, Minas Gerais, Brazil. The 275 blood smears were used both for the microscopical examination and nested PCR providing the DNA template used for the reactions. The sensitivity of the nested PCR assay was higher than that observed for blood smears through microscopical examination. High prevalence (39.6%) of Plasmodium infections was detected by nested PCR while the microscopical examination detected only 16.5 % positive birds. Poor agreement was observed between the results of the two different tests. The PCR data obtained were correlated to the forest fragment size of the Atlantic Forest and also correlated to the biological characteristics of the birds (nest type construction, diet, participation in mixed-species flocks, age and sex). Birds captured in the large forest areas were more infected than birds captured in the small areas (51.9 % and 28.5 %, respectively). Diet and participation in mixed-species flocks were correlated to the Plasmodium parasitism. The insectivorous birds and those that participated in mixed-species flocks were more frequently infected (47% and 41.5%, respectively) than the other groups.

Lutzomyia longipalpis peritrophic matrix: formation, structure, and chemical composition.

Sandflies are vectors of several pathogens, constituting serious health problems. Lutzomyia longipalpis (Lutz & Neiva, 1912) is the main vector of Leishmania chagasi, agent of visceral leishmaniasis. They synthesize a thick bag-like structure that surrounds the bloodmeal, named peritrophic matrix (PM). One of the major roles of PM in blood-fed insects includes protection against ingested pathogens by providing a defensive barrier to their development. We used traditional and modern morphological methods as well as biochemical and immunolabeling tools to define details of the PM structure of the Lu. longipalpis sandfly, including composition, synthesis, and degradation. The kinetics of PM formation and degradation was found to be related to the ingestion and time of digestion of the bloodmeal. The midgut changes its size and morphology after the blood ingestion and during the course of digestion. A striking morphological modification takes place in the midgut epithelium after the stretching caused by the bloodmeal, revealing a population of cells that was not observed in the unfed midgut. The transmission and scanning electron microscopies were used to reveal several morphological aspects of PM formation. The PM looks thicker and well formed 24 h after the bloodmeal. Presence of chitin in the PM was demonstrated by immunolabeling with an alpha-chitin monoclonal antibody. SDS-polyacrylamide gel electrophoresis showed at least five protein bands with molecular masses of 38.7-135 kDa, induced by the protein-free diet. Mouse polyclonal antiserum was produced against PMs induced by protein-free meal and used in Western blotting, which revealed at least three associated proteins.

Variability of the salivary proteins of 20 Brazilian populations of Panstrongylus megistus (Hemiptera: Reduviidae: Triatominae).

The objective of the present study was to study variability in the salivary proteins of 20 Panstrongylus megistus populations from different ecotopes and verify whether this variability influenced the intensity of the response to specific anti-saliva antibodies. Electrophoretic analysis of P. megistus saliva showed a complex protein composition and great interpopulation variability. A higher concentration of bands was observed in the 17-29 kDa region. The phenogram constructed from the electrophoretic profiles of the P. megistus study populations revealed the existence of two main groups. However, there was no evident relationship between these groups and geographical regions, ecotopes or hosts. Saliva inoculated by P. megistus during feeding elicited production of low level of anti-saliva antibodies in rabbit. The homologous and heterologous salivary proteins were recognised by serum of rabbit sensitised with saliva from only one population. Qualitative and quantitative differences were observed for recognised bands in the saliva of all eight populations studied by Western blot analysis. The most recognised bands were those of greatest molecular weight (68.0-97.4 kDa).

Vertical toxoplasmosis in a murine model. Protection after immunization with antigens of Toxoplasma gondii incorporated into liposomes.

Distinct Toxoplasma gondii antigens were entrapped within liposomes and evaluated for their ability to protect Balb/c mice against congenital transmission: soluble tachyzoite antigen (L/STAg), soluble tissue cyst antigen (L/SCAg), soluble tachyzoite plus tissue cyst (L/STCAg) or purified 32kDa antigen of tachyzoite (L/pTAg). Soluble tachyzoite antigen alone in PBS (STAg) or emulsified in Freund's Complete Adjuvant (FCA/STAg) was also evaluated. Dams were inoculated subcutaneously with these antigens 6, 4 and 2 weeks prior to a challenge with four tissue cysts of the P strain of T. gondii orally between 10 and 14 days of pregnancy. Significant diminution differences were observed between the frequency of infected pups born of the dams immunized with the antigens incorporated into liposomes and that of pups born of the dams immunized with antigen emulsified in FCA or non immunized group (p<0.05). There was a significant decrease in the number of pups born dead in the groups L/STAg, L/SCAg and L/pTAg when compared with pups from all other groups (p <0.05). All dams immunized with or without adjuvant showed an antibody response and a proliferation of T-cells. However, no correlation was found between immune response and protection against the challenge.

Persistence of humoral response against sporozoite and blood-stage malaria antigens 7 years after a brief exposure to Plasmodium vivax.

The persistence of malarial antibodies was evaluated in subjects living in a rural community (in Minas Gerais State, Brazil) briefly exposed to a Plasmodium vivax malaria outbreak outside of the area in which malaria was endemic. Transmission was interrupted by treatment of all patients and their relatives and/or neighbors, although the latter had neither symptoms nor blood parasites. Antibodies to P. vivax antigens (recombinant proteins from sporozoites [rPvCS] and from blood stages [rPv200]) were measured in parallel by ELISA with sera collected at two time points after transmission. Anti-rPvCS IgG antibodies were positive in approximately 40% and 20% of the subjects 8 months and 7 years after exposure, respectively. Anti-rPv200 IgG was first detected in 61% of the subjects who had had malarial symptoms and remained positive in 47% after 7 years. Among the prophylactically treated group, anti-rPv200 IgG was detected in only 28% after 8 months. The levels of both antibodies decreased with time in all positive subjects.

Comparison of circumsporozoite proteins from avian and mammalian malarias: biological and phylogenetic implications.

The circumsporozoite (CS) protein of malaria parasites (Plasmodium) covers the surface of sporozoites that invade hepatocytes in mammalian hosts and macrophages in avian hosts. CS genes have been characterized from many Plasmodium that infect mammals; two domains of the corresponding proteins, identified initially by their conservation (region I and region II), have been implicated in binding to hepatocytes. The CS gene from the avian parasite Plasmodium gallinaceum was characterized to compare these functional domains to those of mammalian Plasmodium and for the study of Plasmodium evolution. The P. gallinaceum protein has the characteristics of CS proteins, including a secretory signal sequence, central repeat region, regions of charged amino acids, and an anchor sequence. Comparison with CS signal sequences reveals four distinct groupings, with P. gallinaceum most closely related to the human malaria Plasmodium falciparum. The 5-amino acid sequence designated region I, which is identical in all mammalian CS and implicated in hepatocyte invasion, is different in the avian protein. The P. gallinaceum repeat region consists of 9-amino acid repeats with the consensus sequence QP(A/V)GGNGG(A/V). The conserved motif designated region II-plus, which is associated with targeting the invasion of liver cells, is also conserved in the avian protein. Phylogenetic analysis of the aligned Plasmodium CS sequences yields a tree with a topology similar to the one obtained using sequence data from the small subunit rRNA gene. The phylogeny using the CS gene supports the proposal that the human malaria P. falciparum is significantly more related to avian parasites than to other parasites infecting mammals, although the biology of sporozoite invasion is different between the avian and mammalian species.

Difference in susceptibility to lysis between clones of the Y strain of Trypanosoma cruzi.

Three clones isolated from the Y strain of Trypanosoma cruzi--YP1, YP2 and YP3--were adapted to in vitro cultivation in VERO cells. The recovery of the parasites from the Y strain and clone YP3 was similar after 24 hr of contact with cells (3.2% and 2.7%, respectively) and much lower than the recovery of clones YP1 and YP2 (56.7% and 60.0% of inoculum, respectively). After five days incubation, the ratio Trypomastigotes/Amastigotes released into the supernatants was about 90/10 for clone YP1, YP3 and Y strain, and 50/50 for clone YP2. After nine days, the ratio was 62/38 for clone YP1, 97/3 for clone YP3, 81/19 for Y strain and 50/50 for clone YP2. The susceptibility of tissue culture derived trypomastigotes (TCT) to lysis in the presence of chronic chagasic human sera and human complement was assessed using Complement Mediated Lysis reaction (CML). Trypomastigotes from clone YP2 were consistently less susceptible to CML (% lysis less than 20), than parasites from the other clones and Y strain. Parasites of clone YP3 had susceptibility to CML comparable to that of the Y strain (about 70%), while TCT of clone YP1 had intermediary susceptibility (40%).