Antimicrobial-Resistant Enterococcus spp. in Wild Avifauna from Central Italy.

Bacteria of the genus Enterococcus are opportunistic pathogens, part of the normal intestinal microflora of animals, able to acquire and transfer antimicrobial resistance genes. The aim of this study was to evaluate the possible role of wild avifauna as a source of antimicrobial-resistant enterococci. To assess this purpose, 103 Enterococcus spp. strains were isolated from the feces of wild birds of different species; they were tested for antimicrobial resistance against 21 molecules, vancomycin resistance, and high-level aminoglycosides resistance (HLAR). Furthermore, genes responsible for vancomycin, tetracycline, and HLAR were searched. E. faecium was the most frequently detected species (60.20% of isolates), followed by E. faecalis (34.95% of isolates). Overall, 99.02% of the isolated enterococci were classified as multidrug-resistant, with 19.41% extensively drug-resistant, and 2.91% possible pan drug-resistant strains. Most of the isolates were susceptible to amoxicillin/clavulanic acid (77.67%) and ampicillin (75.73%), with only 5.83% of isolates showing an ampicillin MIC ≥ 64 mg/L. HLAR was detected in 35.92% of isolates, mainly associated with the genes ant(6)-Ia and aac(6')-Ie-aph(2″)-Ia. Few strains (4.85%) were resistant to vancomycin, and the genes vanA and vanB were not detected. A percentage of 54.37% of isolates showed resistance to tetracycline; tet(M) was the most frequently detected gene in these strains. Wild birds may contribute to the spreading of antimicrobial-resistant enterococci, which can affect other animals and humans. Constant monitoring is essential to face up to the evolving antimicrobial resistance issue, and monitoring programs should include wild avifauna, too.

Neuronal satellitosis is a common finding in the avian brain.

Perineuronal or neuronal satellitosis is the term describing the presence of glial cells in the satellite space surrounding the neuronal perikaryon. Confusingly, this finding has been described both as a physiologic and pathologic condition in humans and animals. In animals, neuronal satellitosis has been described in mammals, as well as in avian species. For the latter, the authors wondered whether neuronal satellitosis is expressed in the normal telencephalon of different avian orders and families and whether this pattern in different species shows a specific brain-region association. For these aims, this study explored the presence of neuronal satellitosis in the major areas of the healthy telencephalon in wild and domestic avian species of different orders and families, evaluating its grade in different brain regions. Neuronal satellitosis was seen in the hyperpallium and mesopallium as areas with the highest grade. Passeriformes showed the highest grade of neuronal satellitosis compared to diurnal or nocturnal raptors, and Charadriiformes. To clarify the exact role of neuronal satellitosis in animals without neurological disease, further studies are needed.RESEARCH HIGHLIGHTSNeuronal satellitosis is a common finding in the healthy avian telencephalon.Neuronal satellitosis is a species- and brain-region-associated finding in birds.Passeriformes have the highest grade of neuronal satellitosis.

Molecular Detection of Avipoxvirus in Wild Birds in Central Italy.

Avipoxviruses (APVs) are important pathogens of both domestic and wild birds. The associated disease is characterized by skin proliferative lesions in the cutaneous form or by lesions of the first digestive and respiratory tracts in the diphtheritic form. Previous studies investigated these infections in symptomatic wild birds worldwide, including Italy, but data about the circulation of APVs in healthy avian wildlife are not available. The present study tested spleen samples from 300 wild birds without typical lesions to detect Avipoxvirus DNA. Overall, 43.33% of the samples scored positive. Aquatic birds were more frequently infected (55.42%) than other animals (26.40%), and in Anseriformes, high positivity was found (52.87%). The obtained results suggest that wild birds could be asymptomatic carriers of Avipoxviruses, opening new possible epidemiological scenarios.

Landscape and Climatic Variations Shaped Secondary Contacts amid Barn Owls of the Western Palearctic.

The combined actions of climatic variations and landscape barriers shape the history of natural populations. When organisms follow their shifting niches, obstacles in the landscape can lead to the splitting of populations, on which evolution will then act independently. When two such populations are reunited, secondary contact occurs in a broad range of admixture patterns, from narrow hybrid zones to the complete dissolution of lineages. A previous study suggested that barn owls colonized the Western Palearctic after the last glaciation in a ring-like fashion around the Mediterranean Sea, and conjectured an admixture zone in the Balkans. Here, we take advantage of whole-genome sequences of 94 individuals across the Western Palearctic to reveal the complex history of the species in the region using observational and modeling approaches. Even though our results confirm that two distinct lineages colonized the region, one in Europe and one in the Levant, they suggest that it predates the last glaciation and identify a secondary contact zone between the two in Anatolia. We also show that barn owls recolonized Europe after the glaciation from two distinct glacial refugia: a previously identified western one in Iberia and a new eastern one in Italy. Both glacial lineages now communicate via eastern Europe, in a wide and permeable contact zone. This complex history of populations enlightens the taxonomy of Tyto alba in the region, highlights the key role played by mountain ranges and large water bodies as barriers and illustrates the power of population genomics in uncovering intricate demographic patterns.

Genomic consequences of colonisation, migration and genetic drift in barn owl insular populations of the eastern Mediterranean.

The study of insular populations was key in the development of evolutionary theory. The successful colonisation of an island depends on the geographic context, and specific characteristics of the organism and the island, but also on stochastic processes. As a result, apparently identical islands may harbour populations with contrasting histories. Here, we use whole genome sequences of 65 barn owls to investigate the patterns of inbreeding and genetic diversity of insular populations in the eastern Mediterranean Sea. We focus on Crete and Cyprus, islands with similar size, climate and distance to mainland, that provide natural replicates for a comparative analysis of the impacts of microevolutionary processes on isolated populations. We show that barn owl populations from each island have a separate origin, Crete being genetically more similar to other Greek islands and mainland Greece, and Cyprus more similar to the Levant. Further, our data show that their respective demographic histories following colonisation were also distinct. On the one hand, Crete harbours a small population and maintains very low levels of gene flow with neighbouring populations. This has resulted in low genetic diversity, strong genetic drift, increased relatedness in the population and remote inbreeding. Cyprus, on the other hand, appears to maintain enough gene flow with the mainland to avoid such an outcome. Our study provides a comparative population genomic analysis of the effects of neutral processes on a classical island-mainland model system. It provides empirical evidence for the role of stochastic processes in determining the fate of diverging isolated populations.

Morishitium polonicum as a Cause of Severe Respiratory Disease in Eurasian Blackbirds (Turdus merula) in Central Italy.

Two Eurasian Blackbirds (Turdus merula) from central Italy were found with severe cyclocoelid trematodosis associated with airsacculitis. The birds were submitted with severe respiratory distress; one died shortly after hospitalization, while the second bird was euthanized. At necropsy, a massive presence of cyclocoelid flukes was observed in the coelomic cavity and air sacs of both birds. The air sacs were diffusely opaque, thickened, and covered by scant fibrinous exudate mixed with numerous parasites. Histologically, the air sacs showed diffuse and severe oedema with fibrinous exudate. Diffuse mononucleated and heterophilic infiltration mixed with multiple granulomas contained degenerated trematodes. Morishitium polonicum was identified using morphologic keys and molecular analysis of extracted DNA. Infections caused by M. polonicum are poorly documented in blackbirds and the findings in these birds support the pathogenic role of this trematode as a potential cause of death in blackbirds in Italy. Extended epidemiologic surveys are required to properly assess the potential importance of M. polonicum as a life-threatening pathogen in Blackbird populations.

Natural distemper infection in stone martens (Martes foina): From infection to neutralizing antibodies.

We report an outbreak of canine distemper virus (CDV) among stone martens (Martes foina) in Italy. After being rescued in Northern Italy between April and June 2018, six subjects were kept in a wildlife and exotic animal rescue center in Bologna province. Subjects have been monitored for 15 months in captivity. Within this time-lapse, two subjects died, while among the remaining four, only one showed clinical symptoms referable to distemper. Surviving subjects have been regularly tested for CDV by means of reverse transcriptase-PCR from conjunctival and oropharyngeal swabs for eleven months. The identified viruses belonged to the Wildlife-Europe CDV genetic subgroup. Neutralizing antibodies were detected at the end of the eleven months, when all subjects tested reverse transcriptase-PCR negative. Our findings confirm the circulation of the Wildlife-Europe CDV genetic subgroup (Europe 1/South America 1 lineage) within the Italian wildlife, and improve knowledge on viral infection in stone martens.

Haemoproteus spp. and Leucocytozoon californicus Coinfection in a Merlin (Falco colombarius).

The Leucocytozoon genus comprises numerous widely distributed parasites which have been less investigated than other avian hemoprotozoa. Their occurrence is common, with very variable prevalence values and pathogenicity degrees. Leucocytozoon species are characterized by a great taxonomic diversity, and infections are usually restricted to birds of the same family. In the present paper, a mixed hemosporidia infection by Leucocytozoon californicus and Haemoproteus sp. in an adult male merlin (Falco columbarius) which died during hospitalisation is reported, indicating, for the first time, a newly described avian host species. A molecular investigation was carried out through cytochrome b gene analysis, revealing a 100% match with L. californicus and Haemoproteus spp. A blood smear examination allowed us to detect Leucocytozoon fusiform mature gametocytes and different degrees of maturity of Haemoproteus gametocytes. Histopathology revealed foci of necrosis, hemorrhagic areas and extramedullary hematopoiesis in the liver, the presence of microthrombi in the heart and lung and scattered hemorrhages in the lung.

Effect of topical ophthalmic instillation of rocuronium bromide on the intraocular pressure of kestrels (Falco tinnunculus) and little owls (Athene noctuae).

OBJECTIVE: To assess the effect of rocuronium bromide-induced mydriasis on the intraocular pressure (IOP) of kestrels (Falco tinnunculus) and little owls (Athene noctuae). ANIMALS: 13 adult kestrels and 13 adult little owls. PROCEDURES: All birds were ophthalmologically normal. During the first of 2 treatment periods, a 1% rocuronium bromide solution was topically instilled in both eyes of all birds at a dose of 0.12 mg (12 µL) for kestrels and 0.20 mg (20 µL) for little owls. No ophthalmic treatments were administered during the second (control) treatment period, which was conducted 1 week after the first. During both treatment periods, rebound tonometry was used to measure IOP before rocuronium bromide instillation or at the beginning of the control period (baseline) and at predetermined times after baseline or until the pupillary light reflex returned to normal. All IOP measurements were obtained between 8 am and 5 pm. RESULTS: The mean IOP did not differ significantly from baseline for either species during the control treatment period. During the rocuronium bromide treatment period, complete mydriasis was achieved in all birds. The mean IOP was significantly decreased from baseline and from the corresponding mean IOP for the control period beginning 60 and 30 minutes after drug instillation for kestrels and owls, respectively, and reached its nadir at 60 minutes after drug instillation for both species. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that topical instillation of rocuronium bromide in the eyes successfully induced mydriasis and decreased the IOP of common kestrels and little owls.

Detection of Neospora Caninum DNA in Wild Birds from Italy.

The role of avian species in the Neospora caninum life cycle has not been completely elucidated, and epidemiological data from Europe are scant. The aim of the present report was to evaluate the presence of N. caninum DNA in the tissues of 302 birds belonging to different avian species, along with IFAT titers. Forty-two out of the 302 birds (13.9%) showed low serological titers (1/50 and 1/100) against N. caninum. These data, positive for 31 animals (10.3%), were corroborated by PCR. Twenty-two hearts and eighteen brains scored positive, while nine subjects resulted had parasite DNA both in their hearts and brains. Serological data showed significantly higher results in waterfowl in respect to non-waterfowl avian species. This finding indicates a higher exposure of water birds to the parasite. These avian species, in fact, which feed directly from soil and/or water, are prone to ingest oocysts excreted by final canid hosts. The present study adds information to the state of art of N. caninum epidemiology in Italy, even if more investigations using bio-assays are needed to allow for a serological/parasitological follow-up to evaluate the real impact of the avian species in maintaining the parasite in main reservoirs.

Serological and Molecular Investigation on Toxoplasma gondii Infection in Wild Birds.

Toxoplasma gondii is an obligate apicomplexan zoonotic parasite that infects humans and other animals and is responsible for toxoplasmosis. This parasite causes one of the most common parasitic infections in humans worldwide. Toxoplasmosis meets the requirements for a One Health Disease due to its ability to affect the health of human beings as well as domestic and free ranging animals. Integrating human, domestic animal, and wildlife data could better assess the risk and devise methods of control. A first step of such an approach would be the knowledge of the prevalence of parasitosis in humans and animals in selected areas. Therefore, the aim of the present study was to evaluate the occurrence of Toxoplasma infection in 216 free ranging birds belonging to different genera/species by serology and molecular techniques. Twenty-five out of 216 animals (11.6%) were positive to the immunofluorescence antibody test (IFAT) with antibody titers ranging from 1/20 to 1/320, and 19 of them (8.8%) also showed a positive PCR for Toxoplasma DNA. The results confirmed the widespread occurrence of Toxoplasma infection in wild birds and serological data were corroborated by molecular results in birds that also had low antibody titers. The knowledge of the wide occurrence of the parasite in game and wild birds should enhance the accurate estimation of the risks in handling, managing, and eating these species with regard to domestic carnivores as well as the impact of viscera and offal in the environment.

Morphological study of the iris musculature in diurnal and nocturnal raptors / Estudio morfológico de la musculatura del diafragma en aves rapaces diurnas y nocturnas

In literature it is established that the iris musculature consists of striate muscle fibers in birds while in mammals it consists of smooth muscles. Some authors report the presence of smooth muscle tissue also in the iris of some species of birds. In the present study we report on the iris muscle tissues (type of tissue, direction and mean diameter of muscle fibers or cells) in five species of Accipitriformes (diurnal raptors) and four species of Strigiformes (nocturnal raptors) because they show different way of life depending of their predatory behavior. This morphological study was carried out from raptors died or euthanized at the Wild Life Rescue Centre of Sea and Water birds in Livorno (Italy). From histological examination of iris serial radial sections we find both striated and smooth musculature even if with marked differences among analyzed species, not directly correlated with diurnal or nocturnal lifestyle. Striated fibers are always present, mainly with cross direction, throughout the iris stroma, while the histological differences concern the smooth cells. Indeed, harrier and sparrow hawk (Accipitriformes) and great horned owl and little owl (Strigiformes) show a compact layer of cross smooth muscle cells throughout the iris stroma. In the other species analyzed smooth muscle cells are slightly detectable as scattered or not detectable. Since the cross smooth muscle tissue allows to maintain a myotic state for extended periods of time, our results might be correlated more to the predatory behavior than the taxonomic order.

En la literatura, se establece que la musculatura del iris se compone de fibras musculares estriadas en las aves, mientras que en los mamíferos, la forman los músculos lisos. Algunos autores informan también de la presencia de tejido muscular liso en el iris de algunas especies de aves. El presente estudio informa sobre los tejidos musculares del iris (tipo de tejido, la dirección y diámetro de las fibras musculares o células) en cinco especies de Accipitriformes y cuatro especies de Strigiformes que muestran diferentes hábitos en función de su comportamiento depredador. Este estudio morfológico se realizó en aves rapaces que murieron o fueron eutanasiadas en el Centro de Vida Salvaje de Rescate de Aves Marinas y Acuáticas de Livorno (Italia). El examen histológico de secciones seriadas radiales del iris mostró la presencia tanto de musculatura estriada como lisa, aunque con marcadas diferencias entre las especies analizadas, pero sin correlación directa con el estilo de vida diurna o nocturna. Las fibras estriadas estuvieron siempre presentes, principalmente en dirección transversal a lo largo del estroma del iris, mientras que las diferencias histológicas fueron de las células lisas. Tanto el aguilucho y el gavilán (Accipitriformes) como el buho real y el mochuelo (Strigiformes) mostraron una capa compacta de células musculares lisas transversales en todo el estroma del iris. En las otras especies analizadas, las células musculares lisas fueron ligeramente detectables de manera dispersa o no detectables. Dado que el tejido del músculo liso transversal permite mantener un estado miótico durante largos períodos de tiempo, nuestros resultados podrían estar más correlacionados con el comportamiento depredador, y no con el orden taxonómico.

Schirmer tear test type I readings and intraocular pressure values assessed by applanation tonometry (Tonopen® XL) in normal eyes of four European species of birds of prey.

OBJECTIVE: To determine normal values for Schirmer tear test I and intraocular pressure in four European species of birds of prey. ANIMALS STUDIED: Twenty birds from each of the following species: Eurasian Tawny owl (Strix aluco), Little owl (Athene noctua), Common buzzard (Buteo buteo), and European kestrel (Falco tinnunculus). PROCEDURES: Both eyes of all birds (80 eyes) underwent a complete ophthalmic examination, which included a Schirmer tear test type I (STT-I) performed with commercially available strips and the assessment of the intraocular pressure (IOP) by applanation tonometry, employing the Tonopen-XL(®) device. The animals, which had been taken to a rescue center, were examined for ocular lesions prior to their eventual release into the wild. STT-I readings and IOP values were expressed as means ± standard deviation. RESULTS: Schirmer tear test type I readings were as follows: Eurasian Tawny owls: 3.12 ± 1.92 mm/min; Little owls: 3.5 ± 1.96 mm/min; Common buzzards: 12.47 ± 2.66 mm/min; European kestrels: 6.20 ± 3.67 mm/min. IOP values were as follows: Eurasian Tawny owls: 11.21 ± 3.12 mmHg; Little owls: 9.83 ± 3.41 mmHg; Common buzzards: 17.2 ± 3.53 mmHg; European kestrels: 8.53 ± 1.59 mmHg. CONCLUSIONS: The results of this study give representative values for STT-I and IOP in four of the most common species of birds of prey in Europe.

Safety and efficacy of bilateral topical application of rocuronium bromide for mydriasis in European kestrels (Falco tinnunculus).

The avian iris is mainly composed of striated muscle fibers, and the topical parasympatholytic drugs used in mammals to induce pupil dilation are ineffective in birds. The achievement of mydriasis in conscious birds is only possible by using neuromuscular blocking agents such as curariform drugs. To determine the efficacy of the neuromuscular blocking agent rocuronium bromide as a mydriatic agent in European kestrels (Falco tinnunculus) and to assess possible adverse effects due to its use, 10 kestrels received a single topical instillation with a pipette of 0.12 mg of rocuronium in each eye (total dose, 0.24 mg/bird). Pupil diameter was measured with a pupillary gauge, and pupillary light reflex was assessed by using a standard light source from time zero (Tbase) to 250 minutes after application (T250). Baseline pupil size (Tbase) was 3.30 (0.71) mm in both eyes. Maximal mydriasis was achieved at T90 minutes with a mean (SD) pupillary diameter of 6.30 (0.42) mm and 6.35 (0.41) mm in the right eye and left eye, respectively. At the dose used, rocuronium bromide induced a consistent bilateral simultaneous mydriasis without noticeable adverse effects, and it can be considered a reliable and safe topical mydriatic agent in kestrels.

Bilateral mydriasis in common buzzards (Buteo buteo) and little owls (Athene noctua) induced by concurrent topical administration of rocuronium bromide.

OBJECTIVE: To evaluate the achievement of a bilateral mydriasis in raptors induced by a concurrent topical application of rocuronium bromide and to assess any side effects that might result from its use. Animals studied Ten healthy adult common buzzards (Buteo buteo) and 10 healthy adult little owls (Athene noctua). PROCEDURES: Common buzzards (Group 1) received a single dose of 0.40 mg of rocuronium bromide in each eye (total dose 0.80 mg/bird), whereas the little owls (Group 2) received a single dose of 0.20 mg in each eye (total dose 0.40 mg/bird). The drug was topically instilled in all the birds of both groups. The pupil diameter was measured with a pupillary gauge and the assessment of the pupillary light reflexes was performed using a standard light source. RESULTS: Maximal pupillary diameter was 8.10 ± 0.56 mm in the right eye and 8.05 ± 0.59 mm in the left eye for Group 1 and 10.0 ± 0.75 mm in both eyes for Group 2. No statistical differences were evidenced between the achieved pupillary diameters of both eyes in each group. The maximal pupillary diameter was achieved at T110 min and T40 min for Groups 1 and 2, respectively. The drug did not cause noticeable adverse effects in the examined birds. CONCLUSIONS: A single concurrent topical administration of rocuronium bromide to the eyes of the examined birds induced a complete bilateral mydriasis in both eyes without causing any adverse effect.

Mydriatic effect of topically applied rocuronium bromide in tawny owls (Strix aluco): comparison between two protocols.

OBJECTIVE: To evaluate the mydriatic efficacy of a neuromuscular blocking agent (rocuronium bromide) applied topically to only one eye of nocturnal birds of prey and to assess for any general and/or local adverse effects due to its use. Animal studied Twelve healthy adult tawny owls (Strix aluco) were randomly divided in two groups. PROCEDURES: Six birds (Group 1) received a single dose of 0.35 mg of rocuronium bromide. The second group of subjects (Group 2) received two doses of 0.35 mg of rocuronium bromide (total 0.70 mg/eye). In both groups, the curariform agent was instilled topically. Pupil diameter was measured with a pupillary gauge in 10 min intervals for a total of 100 min and then every 20 min for a total of 240 min. The assessment of the pupillary light reflex was performed using a standard light source during pupillary size recording. RESULTS: Maximal pupillary diameter was 11.5 ± 0.3 mm for Group 1 and 11.0 ± 0.6 mm for Group 2 and no statistically significant differences were detected among the two groups. The maximal pupillary diameter was achieved at T80 for Group 1, and at T60 for Group 2. A complete fundus examination was possible on all treated eyes of subjects of both groups. The drug did not cause any noticeable adverse effects in any of the examined birds. CONCLUSION: Results of the present study suggest that a single topical administration of 0.35 mg of rocuronium bromide to the eyes of healthy tawny owls results in sufficient mydriasis to allow for a complete examination of the fundus.

Aspergillosis in Larus cachinnans micaellis: survey of eight cases.

Avian aspergillosis is reported in several avian species, with Aspergillus fumigatus as the main aetiological agent. Predisposing factors such as starvation, thermal stress, migratory stress, primary infectious disease or toxicosis may play a role. Eight cases of disseminated aspergillosis in free ranging seagulls sheltered at C.R.U.M.A. (Centro Recupero Uccelli Marini e Acquatici, Livorno, Italy) with different clinical histories are presented. The infection was demonstrated by cultural and histological methods from lesions of all birds, and the presence of airborne A. fumigatus viable elements ranging from 450 to 525 CFU/m(3) inside and outside the shelter by means of a surface air sampler (SAS) Super-90 was also assessed. The role of this fungal species as an opportunistic factor in the captivity of seagulls is considered and some control measures, such as a clean and stress free environment and the use of antifungal drugs are suggested.

Sertoli cell tumor associated with polyomavirus infection in a Gouldian finch (Erythrura gouldiae).

A 3-yr-old male Gouldian finch (Erythrura gouldiae) died after 2 wk of lethargy, emaciation, feather loss, and abdominal distension. The bird was housed in an aviary for breeding, but it had shown loss of fertility in the previous breeding season. Necropsy revealed a gross, firm, and yellow mass involving the left testis. Histologically, the mass was a mixed form, intratubular and diffuse, Sertoli cell tumor. Some neoplastic cells had intranuclear inclusion bodies that immunoelectron microscopy proved to be polyomavirus particle aggregates. There were no viral inclusions in other tissues. The possible role of infection in the pathogenesis of the tumor is discussed.

Occurrence of yeasts in psittacines droppings from captive birds in Italy.

Three-hundred twenty five droppings from parrots raised in the premises of 4 breeders and in several private households were cultured for yeasts. One-hundred sixty droppings (49.2%) resulted positive. From these specimens 212 isolates belonging to 27 different species were obtained. Mainly Candida species such as C. albicans, C. catenulata, C. curvata, C. famata, C. glabrata, C. guilliermondi, C. holmii, C. intermedia, C. krusei, C. lambica, C. lusitaniae, C. membranaefaciens, C. parapsilosis, C. pelliculosa, C. sake and C. valida were isolated. Debarvomyces marama, D. polymorphus, Geotrichum sp., Pichia etchelsii, P. ohmeri, Rhodotorula glutinis, R. rubra, Rhodotorula sp., Saccharomyces cerevisiae, S. kluyiveri and Zygosaccharomyces sp. were also obtained. Dark colonies on Staib medium were never observed. The psittacine birds apparently serve as carriers for several Candida species or their perfect states and to a lesser extent for other opportunistic yeasts such as Rhodotorula, Trichosporon and Saccharomyces spp., which are considered part of the transient microbiota of the gastrointestinal tract. The most striking finding was the absence of Cryptococcus spp. among the isolates. The present survey confirms the role of pet birds in carrying potential zoonotic yeasts.