Determining the Prevalence of Avian Chlamydiosis in Wild Amazona Species From Brazil Using Molecular Testing and Clinical Signs.

Avian chlamydiosis is a disease that occurs in birds, especially parrots, and is caused by the Gram-negative bacterium Chlamydia psittaci. Wild Animal Screening Centers in Brazil receive, maintain, treat, and place (preferably to nature) wild animals recovered from illegal trafficking. We performed molecular testing for avian chlamydiosis in parrots from the genus Amazona that were presented to these centers. Cloacal swab samples were collected from 59 parrots (Amazona species) and transported in aqueous or culture medium. The samples were subsequently submitted for DNA extraction by the boiling method, polymerase chain reaction (PCR) amplification using CPF/CPR primers, and agarose gel electrophoresis. Conjunctivitis, nasal discharge, and poor body condition were the clinical signs associated with a differential disease diagnosis of avian chlamydiosis. Transport medium did not have an effect on the test results. The prevalence of C psittaci in the samples was 37% (22/59, 95% confidence interval: 25-49). There was a significant (P = 0.009) association between the PCR test results and clinical signs. Follow-up testing was conducted on a subgroup of 14 individuals that initially tested negative on PCR; 50% (7/14) of these birds were found to be positive within 24 days of the first test. The results of this study confirm the feasibility of using the CPF/CFP primer-based PCR to detect C psittaci in Amazona species, describe a less costly method of transporting biological material for DNA extraction, and evaluate the temporal aspect for obtaining positive results through molecular testing for C psittaci in Amazona species.

The prevalence of Chlamydia psittaci in confiscated Psittacidae in Colombia.

Chlamydia psittaci is a highly zoonotic bacteria distributed worldwide; it is responsible for psittacosis, one of the most important infectious diseases affecting the Psittacidae, mostly parrots. This work was aimed at determining C. psittaci prevalence and genotype in 177 parrots confiscated in Colombia; cloacal swab (166) and faecal (177) samples were analysed from birds confiscated and housed in a Temporary Wildlife Reception Centre (Centro de Reception de Fauna Temporal). Conventional PCR was run on the samples for amplifying the MOMP gene and then the ompA gene. The C. psittaci genotype A was found in 81.3 % (144/177) of the birds analysed. Cloacal swabs accounted for 129/166 (77.7 %) positive samples and faecal matter for 53/177 (29.9 %), 38 birds proving positive for both types of sample; there was an 8.15 times greater probability of detection for cloacal swabs compared to faecal swabs (p < 0.05). Clinical examination findings were correlated with the animals' positivity for cloacal swabs, faecal matter or both, finding a statistically significant relationship with low respiratory rate (p < 0.05) and broken plumage for cloacal swab sample results (p < 0.1). Even though 85 % seroprevalence has previously been reported in Colombia using indirect ELISA, this study reports for the first time C. psittaci genotype A endemicity in psittacines in captivity in Colombia using molecular techniques, considering the zoonotic risk involved in having these birds as pets.

Detection and molecular characterization of Chlamydia psittaci and Chlamydia abortus in psittacine pet birds in Buenos Aires province, Argentina.

In order to determine the presence and genetic diversity of Chlamydia spp. in the north-eastern area of Buenos Aires province, Argentina, conjunctival, oropharyngeal, cloacal swab and tissues were collected from a total of 90 psittacine pet birds of different age and clinical manifestations. Through molecular methods, Chlamydiaceae was detected in 30% (27/90) of the samples, out of which 70.3% (19/27) were positive for Chlamydia psittaci and 14.9% (4/27) for Chlamydia abortus. Nine C. psittaci positive samples were genotyped by ompA gene sequences, 8 clustered within genotype A and 1 within genotype B. A significant association was observed between the presence of Chlamydia spp. and the manifestation of clinical signs compatible with chlamydiosis, as well as with the age of the birds (younger than one year old). This report contributes to the improvement of our understanding of chlamydial agents in our country.

Phylogenetic analyses of Chlamydia psittaci ompA gene sequences from captive blue-fronted Amazon parrots (Amazona aestiva) with hepatic disease in Brazil.

Chlamydia psittaci was detected in 152 (72%) blue-fronted Amazon parrots (Amazona aestiva, parrot from the Psittacidae family) out of a population of 212 that died during 2009-2011 in a wildlife rescue and rehabilitation centre in Minas Gerais, Brazil, following rescue from illegal wildlife trafficking. The macroscopic changes observed in these animals were hepatomegaly with multifocal white foci visible at the serosal surfaces of the liver, and extending into the parenchyma, and splenomegaly. The microscopic lesions observed in the liver included multifocal to coalescing miliary necrosis of hepatocytes with infiltration by heterophils, lymphocytes and plasma cells. In the spleen, loss of the normal architecture and infiltration by macrophages and plasma cells were observed. Stained tissue sections (Gimenez technique) revealed small round clusters suggestive of C. psittaci (reticulate bodies) in the cytoplasm of macrophages from the liver and spleen. Nine sequences of segments of the ompA gene, obtained from different individuals, were randomly selected for sequencing. The phylogenetic analyses showed that all strains clustered with genotype A, which is the most virulent genotype for birds. This genotype is involved in mortality of psittacines, is easily transmitted in captivity and represents a problem for successful rehabilitation. The results indicate the necessity to improve biosecurity in triage and to provide individual personal protection for professionals and caretakers.

Chlamydia psittaci a été détectée chez 152 (72 %) amazones à front bleu (Amazona aestiva, perroquet de la famille des Psittacidés) sur un total de 212 individus rescapés du trafic illégal et décédés en 2009 et 2011 dans un centre de sauvetage et de réhabilitation de la faune sauvage à Minas Gerais (Brésil). Les modifications macroscopiques observées sur ces oiseaux étaient une hépatomégalie avec des foyers blancs multifocaux visibles sur les surfaces séreuses du foie et s'étendant dans le parenchyme, et une splénomégalie. Les lésions microscopiques observées dans le foie comprenaient une nécrose miliaire multifocale à coalescente des hépatocytes avec infiltration d'hétérophiles, de lymphocytes et de plasmocytes. Dans la rate, une perte de l'architecture normale et l'infiltration de macrophages et de plasmocytes ont été observées. La coloration de coupes de tissus (technique de Gimenez) a révélé de petites grappes rondes évoquant C. psittaci (corps réticulés) dans le cytoplasme des macrophages du foie et de la rate. Neuf produits segmentés d'une partie du gène ompA, obtenus de différents individus, ont été sélectionnés de manière aléatoire pour le séquençage. Les analyses phylogénétiques ont montré que toutes les souches se regroupaient dans le génotype A, qui est le plus virulent pour les oiseaux. Ce génotype est responsable de cas de mortalité chez les psittacidés et se transmet facilement en captivité, ce qui représente un risque pour la réussite des opérations de réhabilitation. Au vu de ces résultats, les auteurs soulignent la nécessité d'améliorer la biosécurité lors du tri des animaux dans les centres de soins et de fournir une protection individuelle aux professionnels et aux gardiens.

Se detectó Chlamydia psittaci en 152 (72%) amazonas frentiazules (Amazona aestiva, loro de la familia Psittacidae) de un total de 212 que murieron durante 2009­2011 en un centro de rescate y rehabilitación de fauna silvestre de Minas Gerais, Brasil, tras haber sido rescatadas del tráfico ilegal. Los cambios macroscópicos que se observaron en estos animales fueron hepatomegalia con focos blancos multifocales visibles en las superficies serosas del hígado y que se extendían hacia el parénquima, y esplenomegalia. Las lesiones microscópicas observadas en el hígado consistieron en necrosis miliar multifocal a coalescente de hepatocitos con infiltración de heterófilos, linfocitos y células plasmáticas. En el bazo, se observó pérdida de la arquitectura normal y infiltración de macrófagos y células plasmáticas. Cortes de tejido teñidos (con la técnica de Giménez) revelaron pequeños racimos redondos que sugerían la presencia de C. psittaci (cuerpos reticulados) en el citoplasma de macrófagos del hígado y del bazo. A partir de distintos individuos, se escogieron aleatoriamente nueve segmentos del gen ompA para ser secuenciados. Los análisis filogenéticos mostraron que todas las cepas correspondían al genotipo A, que es el más virulento para las aves. Este genotipo está involucrado en la mortalidad de psitácidas, se transmite fácilmente en cautiverio y supone un riesgo para el éxito de la rehabilitación. Los resultados indican la necesidad de mejorar la bioseguridad en el triaje y de procurar protección personal individual a profesionales y cuidadores.

Epidemiological and molecular characteristics of Chlamydia psittaci from 8 human cases of psittacosis and 4 related birds in Argentina / Características epidemiológicas y moleculares de Chlamydia psittaci provenientes de 8 casos humanos de psitacosis y de 4 aves relacionadas en la Argentina

In Argentina, the epidemiological and molecular characteristics of Chlamydia psittaci infections are still not sufficiently known. A total of 846 respiratory and 10 ocular samples from patients with suspected human psittacosis were tested for C. psittaci from January 2010 to March 2015. Four samples of birds related to these patients were also studied. Forty-eight samples were positive for C. psittaci by a nested PCR. The molecular characterization of twelve C. psittaci PCR-positive samples received in the National Reference Laboratory INEI-ANLIS "Dr. Carlos G. Malbrán", Buenos Aires, Argentina was performed. Eight positive samples from humans and four from birds were genotyped by ompA gene sequencing. C. psittaci genotype A was found in all human samples and in the related birds. This report contributes to our increasing knowledge of the epidemiological and molecular characteristics of C. psittaci to conduct effective surveillance of its zoonotic infections.

En la Argentina, aún no se conocen suficientemente las características epidemiológicas y moleculares de las infecciones por Chlamydia psittaci. Entre enero del 2010 y marzo del 2015 se estudiaron 846 muestras respiratorias y 10 oculares de pacientes con sospecha de psitacosis para la búsqueda de C. psittaci. También se estudiaron 4 muestras de aves relacionadas con estos pacientes. De ese total, 48 muestras fueron positivas para C. psittaci mediante una reacción en cadena de la polimerasa (PCR) anidada. Posteriormente, se realizó en el INEI-ANLIS «Dr. Carlos G. Malbrán¼ la caracterización molecular de 12 muestras positivas para C. psittaci, 8 de humanos y 4 de aves, que fueron genotipificadas por secuenciación del gen ompA. C. psittaci genotipo A se encontró en todas esas muestras. Este informe contribuye a mejorar nuestro conocimiento de las características epidemiológicas y moleculares de C. psittaci para lograr una vigilancia efectiva de la zoonosis que produce.

Epidemiological and molecular characteristics of Chlamydia psittaci from 8 human cases of psittacosis and 4 related birds in Argentina.

In Argentina, the epidemiological and molecular characteristics of Chlamydia psittaci infections are still not sufficiently known. A total of 846 respiratory and 10 ocular samples from patients with suspected human psittacosis were tested for C. psittaci from January 2010 to March 2015. Four samples of birds related to these patients were also studied. Forty-eight samples were positive for C. psittaci by a nested PCR. The molecular characterization of twelve C. psittaci PCR-positive samples received in the National Reference Laboratory INEI-ANLIS "Dr. Carlos G. Malbrán", Buenos Aires, Argentina was performed. Eight positive samples from humans and four from birds were genotyped by ompA gene sequencing. C. psittaci genotype A was found in all human samples and in the related birds. This report contributes to our increasing knowledge of the epidemiological and molecular characteristics of C. psittaci to conduct effective surveillance of its zoonotic infections.

First Identification of Chlamydia psittaci in the Acute Illness and Death of Endemic and Endangered Psittacine Birds in Mexico.

A mortality episode of endemic and endangered psittacine birds from the genera Ara and Amazona occurred during January 2015. The birds were housed in a management unit for wildlife conservation that receives wild-caught birds from illegal trade. In total, 11 (57%) adult birds of different origins that shared these accommodations died. Only four of them were sent for diagnosis. The main lesions found at necropsy were consistent with those described previously for avian chlamydiosis; the presence of Chlamydiaceae was confirmed through immunofluorescence and amplification with further sequencing of the 16S ribosomal RNA gene by using hepatic tissue. Due to the lack of specific diagnostic tools on primary psittacine diseases, the pathogenic effects of systemic, respiratory, or enteric infections with high mortality rates remain unknown in Mexico. In this study, specific molecular identification of avian chlamydiosis was performed using a nested PCR on liver tissues, as well as choanal and cloacal swab samples, confirming the presence of Chlamydia psittaci in all of them. In addition, it was possible to obtain the ompA gene sequence from processed clinical samples, thereby allowing us to determine that the A genotype was affecting these birds. Although this genotype is the most commonly found worldwide in psittacine birds, this case report describes the first avian chlamydiosis outbreak affecting critically endangered and endemic psittacines subjected to reintegration programs in Mexico. Consequently, this study demonstrates the necessity of more exhaustive biosecurity strategies because other pathogens may be present and should be assessed, especially in highly threatened birds, before releasing them into their habitats.

Risk factors associated with Chlamydia psittaci infection in psittacine birds.

Chlamydia psittaci is the aetiological agent of chlamydiosis in birds, especially Psittaciformes. The objective of the present study was to detect C. psittaci by means of semi-nested PCR among psittacine birds sold at pet markets and kept as pet birds in Salvador, Bahia, Brazil. Questionnaires were used to identify risk factors involved in the epidemiology of the disease. In addition, the management of birds and cages was observed at each location studied. The frequency of C. psittaci infection was 10.6% (33/311) in the psittacine birds studied. Birds kept in households were less frequently positive (3.4%; 5/148) than those at pet markets (17.2%; 28/163). Among the several factors analysed in the epidemiology of the disease, only population density (P = 0.001) and cage hygiene (P = 0.041) in birds at pet markets were significantly associated with C. psittaci infection. These results demonstrate the presence of C. psittaci infection in Psittaciformes kept as pets and held at pet markets in Salvador, Bahia, showing that this micro-organism is a public health concern. Control measures should be encouraged to prevent the spread of the agent among birds, as well as among employees and customers.

Evidence of zoonotic Chlamydophila psittaci transmission in a population at risk in Zulia state, Venezuela.

OBJECTIVE: This study was aimed at investigating the frequency of infection by Cp. psittaci and determining its genotype in individuals at potential risk of exposure to the bacteria. METHODOLOGY: The study involved 170 individuals: a risk group (n= 96) and a low-risk control group (n=74). Cp. psittaci was detected and genotyped by single-tube nested PCR and ompA gene sequencing. RESULTS: Eight (8.3 %) positive cases were detected in the risk group and 1 (1.4 %) in the control group (p<0.04). Cp. psittaci was found in 16.7 % of pigeons' fecal samples. Cp. psittaci infection with was more frequent in symptomatic (17.7 %) than asymptomatic (6.3 %) individuals in the risk group. Analysing the genomes isolated from human and bird specimens revealed the presence of genotype B. CONCLUSION: The presence of Cp. psittaci genotype B in the population being evaluated could have been attributed to zoonotic transmission from pigeons to humans, an underestimated potential public health problem in Venezuela requiring the health authorities' involvement.

Genotyping of C. psittaci in central area of Argentina.

In central area of Argentina, the epidemiologic and molecular characteristics of Chlamydophila psittaci infections are still unknown. Nested polymerase chain reaction of domains II, III, and IV of the omp A gene was used to detect Chlamydophila in 43 pharyngeal swab samples from patients with suspected human psittacosis (2010-2011); 9 (21%) of them yielded positive results. Molecular typing was performed by direct sequencing demonstrating the presence of C. psittaci genotypes A, E/B, and WC.

Evidence of zoonotic Chlamydophila psittaci transmission in a population at risk in Zulia state, Venezuela / Evidencia de transmisión zoonótica de Chlamydophila psittacci en una población de riesgo del Estado Zulia, Venezuela

Objective This study was aimed at investigating the frequency of infection by Cp. psittaci and determining its genotype in individuals at potential risk of exposure to the bacteria. Methodology The study involved 170 individuals: a risk group (n= 96) and a low-risk control group (n=74). Cp. psittaci was detected and genotyped by single-tube nested PCR and ompA gene sequencing. Results Eight (8.3 %) positive cases were detected in the risk group and 1 (1.4 %) in the control group (p<0.04). Cp. psittaci was found in 16.7 % of pigeons' fecal samples. Cp. psittaci infection with was more frequent in symptomatic (17.7 %) than asymptomatic (6.3 %) individuals in the risk group. Analysing the genomes isolated from human and bird specimens revealed the presence of genotype B. Conclusion The presence of Cp. psittaci genotype B in the population being evaluated could have been attributed to zoonotic transmission from pigeons to humans, an underestimated potential public health problem in Venezuela requiring the health authorities' involvement.

Objetivo El objetivo de este estudio fue investigar la frecuencia de infecciones por Cp. psittaci y determinar su genotipo en individuos con potencial riesgo de exposición a la bacteria. Metodología Se incluyeron 170 individuos, un grupo de riesgo (n=96) y un grupo control (n=74). La detección y genotipificación de Cp. psittaci se llevó a cabo por PCR anidada y secuenciación del gen ompA. Resultados Se detectaron ocho (8,3 %) casos positivos en el grupo de riesgo y 1 (1,35 %) en el grupo control (p<0,04). Cp. psittaci fue detectada en 16,7 % muestras fecales de palomas. En el grupo de riesgo, la frecuencia de infección por Cp. psittaci fue 17,7 % en individuos sintomáticos y 6,3% en asintomáticos. El análisis de los genomas aislados de muestras humanas y aves, revelaron la presencia del genotipo B. Conclusión La presencia de Cp. psittaci genotipo B en la población evaluada podría ser atribuida a transmisión zoonótica de palomas a humanos, un potencial problema de salud pública en nuestra región que requiere la intervención de autoridades sanitarias.

Low prevalence of Chlamydia psittaci in ocular adnexal lymphomas from Cuban patients.

Most ocular adnexal lymphomas (OAL) are extranodal marginal zone B-cell lymphomas (EMZL) of mucosa-associated lymphoid tissue (MALT)-type. Chronic antigen stimulation has been suggested to have a pathogenetic role in EMZL and Chlamydia psittaci chronic infection has been recently associated with the development of OAL in a series of patients from Italy. To assess this association, an evaluation of the presence of C. psittaci was made in a different OAL population. DNA samples were obtained from formalin-fixed, paraffin-embedded sections samples of 26 patients with OAL, 20 non-OAL and 20 benign ocular lesions, diagnosed and treated between 1998 and 2003 at National Institute of Oncology in Havana, Cuba. All samples were histologically reviewed by an expert pathologist. Fluorescence in situ hybrization (FISH) analysis of translocations involving MALT1 was performed. The presence of bacterial DNA was assessed with a multiplex touchdown enzyme time release polymerase chain reaction. DNA sequencing was performed to confirm suspicious bands. Seventy-three percent of the OAL cases were EMZL and 81% were in stage IE. FISH analysis was performed in 13 OAL cases and none of them evidenced MALT1 translocations. DNA of C. psittaci was detected in 11% of the 46 lymphomas: two orbital EMZL and three non-OAL. All 20 benign ocular lesions were negative for C. psittaci. The low prevalence of C. psittaci in OAL suggests geographical differences in the etiology of this entity. International studies are needed to clarify the role of C. psittaci in OALs.

[Psittacosis outbreak in Granada, Spain]. / Brote de psitacosis en Granada.

BACKGROUND: Psittacosis can be transmitted to humans from infected birds, being the cause of atypical pneumonia. This work is aimed at determining the source of a psittacosis outbreak, the characteristics of the subjects exposed and describing the measures to control the outbreak. METHODS: Descriptive study. Notification to the Epidemiological Monitoring System and Autonomic Agriculture authorities, taking the pertinent measures (inspection, sampling, disinfecting and precautionary quarantine of birds), closing the establishment. Case definition and epidemiological survey. Frequency analysis. RESULTS: Seventeen (17) individuals (one of whom died) came into contact with suspicious animals (53% males/47% females), nine of whom (mean age: 30) showed symptoms (100% fever, flu-like symptoms 44.4%). A serology was performed for C. psitacci on 11 of the 17 subjects exposed (68.75%), two of whom (18.18%) tested positive. The presence of C. psittaci DNA was detected in the pulmonary tissue study conducted on the deceased individual. A total of 33% (3/9) of those showing symptoms were confirmed by diagnosis. The Autonomic Agriculture authorities placed 22 animal compounds under quarantine. A total of seven (10%) of the 70 samples taken (60: animal cloacas and 10 birds) revealed a positive antigen, the PCR nucleic acid test being positive in one there-of. CONCLUSIONS: The exposure to diseased animals at the establishments involved caused the outbreak, the common source of contagion being of an occupational nature. The appropriate coordination/collaboration between the local Health and Agriculture authorities made it possible to prevent any further cases from arising.