Morphobiometry of the scleral ossicle rings in Chelonia mydas sea turtles.

OBJECTIVE: Scleral ossicle rings of reptiles have endoskeletal functions that are not completely understood. Moreover, descriptive reports on the anatomy of those rings are scarce. We tried to make an anatomical description that could contribute to a better understanding of their functions. ANIMAL STUDIED AND PROCEDURES: We quantified, histologically characterized and evaluated the morphobiometry of the scleral ossicles, and measured the aditus orbitae of 25 sea turtle (Chelonia mydas) heads. RESULTS: The aditus orbitae represented about one-third of the total head length and the mean area of the internal opening of each ring was up to 8.37% of the aditus orbitae area. The mean internal diameter of the rings (6.32 mm) was characteristic of scotopic species and the most frequent number of ossicles per ring varied between 11 and 12. Two new classifications were proposed for the ossicle types: plus-Verzahnung (+V) and minus-Verzahnung (-V). The bone tissue revealed a lamellar arrangement typical of compact and resistant bones. CONCLUSION: The obtained data may be used to support and expand the understanding of functions, animal activity patterns, distinctions between taxa and taphonomic interpretations.

An eye on the Shih Tzu dog: Ophthalmic examination findings and ocular surface diagnostics.

OBJECTIVE: To characterize the ocular surface parameters and determine the prevalence of ocular pathology in Shih Tzu dogs. ANIMAL STUDIED: Fifty Shih Tzu dogs (28 male, 22 female). PROCEDURES: Each dog underwent a complete ophthalmic examination (recording any pathology) and a series of diagnostics, allowing for a 10 min-interval between tests: intraocular pressure (IOP), blink rate, palpebral fissure length (PFL), corneal tactile sensation (CTS), Schirmer tear test and nasolacrimal reflex without (STT-1, NL-STT1) and with topical anesthesia (STT-2, NL-STT2), tear ferning, strip meniscometry test (SMT), tear film breakup time (TFBUT), and punctate fluorescein staining (PFS) of the cornea. RESULTS: Mean ± SD test values were as follows: IOP (17.9 ± 3.7 mmHg), blink rate (2.4 ± 1.4 blinks/min), PFL (23.8 ± 1.8 mm), CTS (1.8 ± 0.7 cm), STT-1 (22.0 ± 5.5 mm/min), NL-STT1 (24.2 ± 4.7 mm/min), STT-2 (16.9 ± 6.5 mm/min), NL-STT2 (18.5 ± 7.5 mm/min), SMT (7.5 ± 3.5 mm/5 s), TFBUT (5.3 ± 2.4 s), tear ferning (1.3 ± 0.7), and PFS (1.6 ± 0.6). PFL was significantly greater in male vs. female Shih Tzus (p< .001). Age was negatively correlated with TFBUT results (r = -0.31, p = .027). Lagophthalmos was observed in 82% eyes. Ocular surface pathology was common, including adnexal abnormalities (100% eyes with caruncular trichiasis and medial lower lid entropion) and corneal opacification (27% pigmentation, 20% fibrosis, 12% neovascularization). CONCLUSIONS: Qualitative tear film deficiency (low TFBUT), along with several anatomical abnormalities that promote ocular irritation and reduce globe protection, together help explain the concerningly high prevalence of ocular surface disease in the Shih Tzu breed. Prophylactic measures (e.g., medial canthoplasty, topical lubrication) could be considered to improve ocular health in Shih Tzus.

Imprinting fidelity in mouse iPSCs depends on sex of donor cell and medium formulation.

Reprogramming of somatic cells into induced Pluripotent Stem Cells (iPSCs) is a major leap towards personalised approaches to disease modelling and cell-replacement therapies. However, we still lack the ability to fully control the epigenetic status of iPSCs, which is a major hurdle for their downstream applications. Epigenetic fidelity can be tracked by genomic imprinting, a phenomenon dependent on DNA methylation, which is frequently perturbed in iPSCs by yet unknown reasons. To try to understand the causes underlying these defects, we conducted a thorough imprinting analysis using IMPLICON, a high-throughput method measuring DNA methylation levels, in multiple female and male murine iPSC lines generated under different experimental conditions. Our results show that imprinting defects are remarkably common in iPSCs, but their nature depends on the sex of donor cells and their response to culture conditions. Imprints in female iPSCs resist the initial genome-wide DNA demethylation wave during reprogramming, but ultimately cells accumulate hypomethylation defects irrespective of culture medium formulations. In contrast, imprinting defects on male iPSCs depends on the experimental conditions and arise during reprogramming, being mitigated by the addition of vitamin C (VitC). Our findings are fundamental to further optimise reprogramming strategies and generate iPSCs with a stable epigenome.

Generation and characterization of induced pluripotent stem cell line (IBBISTi004-A) from an Angelman syndrome patient carrying a class II deletion of the maternal chromosome 15q11.2-q13.

Angelman Syndrome is a rare neurodevelopmental disorder caused by several (epi)genetic alterations. The patients present strong neurological impairment due to the absence of a functional maternal UBE3A gene in neurons. Here, we generated and characterized a new induced pluripotent stem cell (iPSC) line from a female child with Angelman syndrome harbouring a class II deletion. iPSCs were reprogrammed from fibroblasts using Sendai viruses. The new iPSCs express pluripotency markers, are capable of trilineage in vitro differentiation and have the expected imprinting status of Angelman syndrome. These iPSCs are a valuable tool to elucidate the pathophysiological mechanisms associated with this disease.

TonoVet Plus®: Higher reliability and repeatability compared with Tono-Pen XL™ and TonoVet® in rabbits.

OBJECTIVE: To evaluate the accuracy and repeatability of the Tono-Pen XL™, TonoVet® and TonoVet Plus® tonometers by manometric evaluation, and to establish adjustment equations for intraocular pressure (IOP) estimates in rabbits. ANIMAL STUDIED: Rabbits. PROCEDURES: A postmortem study was conducted on seven rabbit eyes to verify the correlation between manometry and tonometry with an artificial incremental increase in IOP from 5 and 60 mmHg. A clinical study was conducted to evaluate accuracy and to establish reference values for the species, with measurement of IOP in 17 animals, for 2 consecutive days, with the same tonometers and calibrations used in the postmortem evaluations. RESULTS: There were strong linear trends for all evaluated tonometers. In the in-vivo evaluation, the mean IOP values were: 14.23 ± 1.75 (Tono-Pen XL™); 13.89 ± 2.07 (TonoVet® calibration mode 'd'); 8.88 ± 1.24 (TonoVet calibration mode 'p'); 18.59 ± 1.94 (Tonovet Plus®). There was a significant difference in the two evaluation times for the two TonoVet® calibration modes. The adjustment equations generated from the manometry for the evaluated tonometers were: Y = 0.2570X + 2.219 (Tono-Pen XL™), Y = 0.2289X + 2.389 (TonoVet® 'd'), Y = 0.4043X + 4.062 (TonoVet® 'p'), Y = 0.1233X + 0.3644 (TonoVet Plus®) (X is device-estimated IOP). CONCLUSIONS: All evaluated tonometers were well correlated with the manometry, with an underestimation of IOP by all devices. Applying adjustment formulas may compensate for systematic errors. TonoVet Plus® was well tolerated, and showed better repeatability and reliability in successive evaluations.

Birefringence Analyses Reveal Differences in Supramolecular Characteristics of Corneal Stromal Collagen Fibrils Between Falconiformes and Strigiformes.

This study aimed to assess the birefringent properties of corneal stromal collagen fibrils in birds of the orders Falconiformes (diurnal) and Strigiformes (predominantly nocturnal) to compare their supramolecular organizations. In total, 22 corneas of Falconiformes (Caracara plancus, n = 8; Rupornis magnirostris, n = 10; and Falco sparverius, n = 4) and 28 of Strigiformes (Tyto furcata, n = 16; Pseudoscops clamator, n = 6; and Athene cunicularia, n = 6) were processed histotechnically into 8-µm thick sections. Corneal optical retardation (OR) values related to the form and intrinsic fractions of the total birefringence of collagen fibrils were measured using a polarized light microscope equipped with phase compensators. In addition, the coherence coefficients that inform the local orientation of the fibrils were calculated through video image analysis. All assessments were conducted both in the anterior and posterior stroma of the cornea. Differences were significant when P < 0.05. The results showed supraorganizational differences between fibrils in the anterior stroma of Falconiformes and Strigiformes. The OR values were greater (P < 0.0001) for Falconiformes, indicating that the corneas of these birds contain more collagen fibrils or more aggregated collagen fibrils. In contrast, the coherence coefficients were higher (P = 0.016) for Strigiformes, indicating that the corneal collagen fibers in these birds are highly aligned and have few undulations. A multivariate data matrix constructed for Euclidean distance calculations showed that the dissimilarity between Falconiformes and Strigiformes corneas, in terms of the supraorganization of stromal collagen fibrils, was 4.56%. In conclusion, it is possible that the supraorganizational differences reported in this study may be sources of variation in the visual quality of Falconiformes and Strigiformes. This study provides the necessary evidence to encourage further research associating corneal optical performance to supramolecular characteristics of corneal stromal collagen.

The tandem repeat modules of Xist lncRNA: a swiss army knife for the control of X-chromosome inactivation.

X-inactive-specific transcript (Xist) is a long non-coding RNA (lncRNA) essential for X-chromosome inactivation (XCI) in female placental mammals. Thirty years after its discovery, it is still puzzling how this lncRNA triggers major structural and transcriptional changes leading to the stable silencing of an entire chromosome. Recently, a series of studies in mouse cells have uncovered domains of functional specialization within Xist mapping to conserved tandem repeat regions, known as Repeats A-to-F. These functional domains interact with various RNA binding proteins (RBPs) and fold into distinct RNA structures to execute specific tasks in a synergistic and coordinated manner during the inactivation process. This modular organization of Xist is mostly conserved in humans, but recent data point towards differences regarding functional specialization of the tandem repeats between the two species. In this review, we summarize the recent progress on understanding the role of Xist repetitive blocks and their involvement in the molecular mechanisms underlying XCI. We also discuss these findings in the light of the similarities and differences between mouse and human Xist.

The glycoproteomics of hawk and caiman tears.

BACKGROUND: Glycoproteins are important tear components that participate in the stability of the ocular surface. However, the glycopeptides that are present in the tears of wild animals have not yet been described. This work aimed to describe the glycoproteomic profile of roadside hawk (Rupornis magnirostris) and caiman (Caiman latirostris) tears. METHODS: Tears collected from 10 hawks and 70 caimans using Schirmer tear test strips were used in this study. The samples were submitted to trypsin digestion and separated using a reverse-phase column coupled to a mass spectrometer associated to a nanospray ionization source. The glycoproteins were categorized as: cellular components, biological processes and molecular function, according to the UniProt Knowledgebase. RESULTS: As shown by the liquid chromatography-mass spectrometry, all glycopeptides found were classified as N-type. Of the 51 glycoproteins that were identified in the hawk tear film, the most abundant were ovotransferrin, globulins and complement system proteins. In the caiman tear film, 29 glycoproteins were identified. The most abundant caiman glycoproteins were uncharacterized proteins, ATPases, globulins and proteasome components. Ontological characterization revealed that the glycoproteins were extracellular, and the most identified molecular function was endopeptidase activity for both species. CONCLUSION: Glycoproteins are abundant in the tear film of the bird and reptile species studied herein, and all these molecules were shown to have N-type modifications. Location at the extracellular space and an endopeptidase inhibitor activity were the main cell component and molecular function for both species, respectively. These profiles showed differences when compared to human tears, are possibly linked to adaptive processes and can be the basis for further studies on the search of disease biomarkers.

Generation and characterization of induced pluripotent stem cells heterozygous for the Portuguese BRCA2 founder mutation.

Women who inherit heterozygous mutations in the BRCA2 gene have an increased risk of developing cancer, mainly breast and ovarian tumors. A particular BRCA2 mutation (c.156_157insAlu) is exclusively found in families of Portuguese ancestry and is present in approximately 30% of all Portuguese families with hereditary breast and ovarian cancers. We report the generation and characterization of the first iPSC line from a female donor harboring the Portuguese BRCA2 founder mutation. Skin fibroblasts were reprogrammed using a non-integrative Sendai virus. These iPSCs are a valuable tool to study the origin of BRCA2-associated cancer in its earliest phases.

Generation and characterization of induced pluripotent stem cells from a family carrying the BRCA1 mutation c.3612delA.

How BRCA1 germline mutations predispose to cancer remains poorly understood. Induced pluripotent stem cells (iPSCs) represent an emerging model to investigate the molecular mechanisms underlying malignant transformation in primary cells from individuals who are carriers of deleterious mutations in the BRCA1 gene. Here we report the generation and characterization of iPSC lines from a female donor harboring a germline c.3612delA mutation in the BRCA1 gene and her daughter who does not carry the mutation. Skin fibroblasts were reprogrammed using non-integrative Sendai virus and characterized for their pluripotent properties. These iPSCs are a valuable cellular model for personalized pre-clinical research in the context of BRCA1 mutant hereditary cancers.

H4K20me1 and H3K27me3 are concurrently loaded onto the inactive X chromosome but dispensable for inducing gene silencing.

During X chromosome inactivation (XCI), in female placental mammals, gene silencing is initiated by the Xist long non-coding RNA. Xist accumulation at the X leads to enrichment of specific chromatin marks, including PRC2-dependent H3K27me3 and SETD8-dependent H4K20me1. However, the dynamics of this process in relation to Xist RNA accumulation remains unknown as is the involvement of H4K20me1 in initiating gene silencing. To follow XCI dynamics in living cells, we developed a genetically encoded, H3K27me3-specific intracellular antibody or H3K27me3-mintbody. By combining live-cell imaging of H3K27me3, H4K20me1, the X chromosome and Xist RNA, with ChIP-seq analysis we uncover concurrent accumulation of both marks during XCI, albeit with distinct genomic distributions. Furthermore, using a Xist B and C repeat mutant, which still shows gene silencing on the X but not H3K27me3 deposition, we also find a complete lack of H4K20me1 enrichment. This demonstrates that H4K20me1 is dispensable for the initiation of gene silencing, although it may have a role in the chromatin compaction that characterises facultative heterochromatin.

Atypical ocular Chelonoid herpesvirus manifestations in a captive Loggerhead turtle (Caretta caretta).

A captive loggerhead turtle (Caretta caretta) of unknown sex, 3 years of age, presented with bilateral mucoid secretions, severe chemosis, conjunctival hyperemia, and globe retraction. The animal was evaluated ophthalmologically and systemically, and hematological, microbiological, and conjunctival cytological and biopsy samples were collected for complementary diagnosis. The histopathological examination showed amphophilic intranuclear inclusions associated with severe inflammatory infiltrate. The diagnosis of Chelonid alphaherpesvirus 5 (ChAHV 5) was confirmed with end point PCR. Following systemic treatment with L-lysine, acyclovir and vitamin A, the ocular signs resolved. No amphophilic intranuclear inclusions were seen in a follow-up biopsy 5 months later, and there has been no recurrence of clinical ophthalmic signs during a 4-year follow-up. It is suggested that ChAHV 5 be considered as a differential diagnosis in captive marine turtles that present for conjunctival disease other than fibropapillomatosis.

Ocular ultrasonography of sea turtles.

BACKGROUND: Environmental changes contribute to the development of ophthalmic diseases in sea turtles, but information on their eye biometrics is scarce. The aim of this study was to describe ophthalmic ultrasonographic features of four different sea turtle species; Caretta caretta (Loggerhead turtle; n = 10), Chelonia mydas (Green turtle; n = 8), Eretmochelys imbricata (Hawksbill turtle; n = 8) and Lepidochelys olivacea (Olive ridley; n = 6) under human care. Corneal thickness, scleral ossicle width and thickness, anterior chamber depth, axial length of the lens, vitreous chamber depth and axial globe length were measured by B-mode sonography with a linear transducer. Carapace size and animal weight were recorded. A sonographic description of the eye structures was established. RESULTS: The four species presented an ovate eyeball, a relatively thin cornea, and a small-sized lens positioned rostrally in the eye bulb, near the cornea, resulting in a shallow anterior chamber. The scleral ossicles did not prevent the evaluation of intraocular structures, even with a rotated eye or closed eyelids; image formation beyond the ossicles and measurements of all proposed structures were possible. B-mode sonography was easily performed in all animals studied. The sonographic characteristics of the eye were similar among the four species. Since there was a correlation between the size of the eye structures and the size of the individual, especially its carapace size, the differences found between E. imbricata and Caretta caretta are believed to be due to their overall difference in size. CONCLUSIONS: Sonography is a valuable tool in ophthalmic evaluation of these species. Only minor differences were found between the species in this study, reinforcing their phylogenetic proximity and their similar functions and habitats.

Comparison of Electrolyte Composition and Crystallization Patterns in Bird and Reptile Tears.

To compare tear electrolytes and tear crystallization patterns in birds and reptiles, tears were sampled by Schirmer tear test from 10 animals each of Ara ararauna, Amazona aestiva, Tyto alba, Rupornis magnirostris, Chelonoidis carbonaria, and Caiman latirostris, and 5 of Caretta caretta. The aliquots were pooled to assess concentrations of total protein, chloride, phosphorus, iron, sodium, potassium, calcium, and urea. For the tear ferning test, samples of each species were observed under a polarized light microscope at room temperature and humidity. Crystallization patterns were graded according Rolando and Masmali scales. There was more total protein and urea in owl and sea turtle tears, respectively, than in the other animals tested. Electrolyte balance was similar for all species, with higher sodium, chloride, and iron. In birds, Rolando-scale grades of tear crystallization patterns ranged from I to II, and from 0 to 2 using the Masmali scale; in reptiles, grades were II to IV (Rolando) and 2 to 4 (Masmali). Crystallization arrangements of some species had higher scores, as caimans and sea turtles, possibly due to different the tear composition. Marine and lacustrine species presented higher. The ionic balance of lacrimal fluids of birds and reptiles was similar to that in humans, with higher values of sodium and chloride. However, a similar tear composition did not influence the crystal morphology. Crystallization classification suggested that higher grades and types are due to the different microelements present in the tears of wild species.

Comparative Analysis of Tear Composition in Humans, Domestic Mammals, Reptiles, and Birds.

Tears are an important component of the ocular surface protection mechanism and are in close contact with the corneal epithelium and the environment. Their composition is well-known in humans; however, there are few investigations on the composition and function of tears in reptiles, birds and others mammals, which would elucidate the mechanisms governing the maintenance of ocular homeostasis. In this work, electrophoretic profiles and an evaluation of total protein, albumin, urea, glucose, and cholesterol concentrations in tears of semi-aquatic, terrestrial, and marine reptiles (Caiman latirostris, Chelonia mydas, Caretta caretta, Eretmochelys imbricata, Lepidochelys olivacea, and Chelonoidis carbonaria), birds (Tyto furcata, Rupornis magnirostris and Ara ararauna), and mammals (Equus caballus and Canis lupus familiaris) were apresented. Human tear components and respective blood serum samples were used as references. The electrophoretic analysis revealed similarities whithin same Classes. The results of the tear-blood serum relationship and the comparison to human tear components showed particularities that are potentially derived from a homeostatic response to the environment. When the tear compositions of animals belonging to different ecological clusters were compared, marked differences were observed in total protein and urea concentrations. Thus, reptile, bird, and mammalian tears are complex fluids with differing concentrations of biochemical components that are potentially a result of the animals' adaptation to different environments.

The use of tear ferning test in cats for evaluation of ocular surface.

BACKGROUND: The tear film is a trilaminar fluid composed mainly of lipids, electrolytes, proteins and water. It is responsible for lubrication, nutrition and protection against microbial and toxic agents. Disruption of any these components may weaken the ocular surface, making it more susceptible to disease. Increasing evidence suggests that qualitative tear film deficiencies are an important predisposing factor or cause of some of the most common and challenging ocular diseases in cats, including conjunctivitis, corneal ulcer, spontaneous chronic corneal epithelial defects (SCCED), pigmentary keratitis, corneal sequestrum and dry eye syndrome. The aim of this study was to describe the tear ferning test in healthy cats and to compare the results by using two grading scales for humans. Tear samples were collected using Schirmer tear test (STT) strips from 60 healthy cats, and, after centrifuging the strips to obtain the samples, the aliquot was placed on clean microscope glass until it dried and the tear ferning patterns were observed under a polarized light microscope and classified according to the Rolando and Masmali grading scales. RESULTS: Ferning patterns in the lower grades showed full crystallization with high density, without gaps between the ferns and branches, forming several nuclei that were easily distinguished. According to the Rolando scale, 50% (60/120), 46.6% (56/120) and 3.4% (4/120) of eyes showed type I, II and III patterns, respectively. According to the Masmali scale, 15% (18/120), 56.6% (68/120 eyes) and 28.4% (34/120) of eyes showed grade 0, 1 and 2 patterns, respectively. No difference was observed between the right and left eyes for both Rolando (P = 0.225) and Masmali (P = 0.683) scales. CONCLUSIONS: The tear ferning test is a qualitative test that can be used in cats as a complementary evaluation of the ocular surface. While the Rolando scale showed an increased prevalence of types I and II, the Masmali scale showed an increased prevalence of grades 1 and 2. This can be attributed to the species-specific differences between human and feline tear film. So Masmali grade 2 can be considered a normal tear pattern for the species, because all the cats used in study were clinically healthy. For this reason, future complementary studies are necessary for comparing healthy eyes and eyes with different ocular surface disease in cats. Both scales can be feasible options for grading tear crystallization in cats, but as Rolando scale included 96.6% of the samples in the 2 types that are considered normal for humans, we think that this scale seemed to be more precise to classify crystallization pattern in cats. The crystallization patterns observed in this study can form the basis for standardizing ocular surface parameters in cats.

Aqueous tear assessment in dogs: Impact of cephalic conformation, inter-test correlations, and test-retest repeatability.

OBJECTIVE: To characterize diagnostic findings, test-retest repeatability, and correlations among lacrimal tests in dogs of diverse cephalic conformations. ANIMAL STUDIED: Fifty healthy dogs (25 brachycephalic, 25 nonbrachycephalic). PROCEDURES: A series of diagnostics were performed in each dog, allowing for a 10-minute interval between tests and repeating each test 24 hours later under similar conditions: corneal tactile sensation (CTS), strip meniscometry test (SMT), phenol red thread test (PRTT), endodontic absorbent paper point tear test (EAPPTT), Schirmer tear test-1 without (STT-1) or with nasolacrimal stimulation (NL-STT1), and Schirmer tear test-2 (STT-2). RESULTS: Mean ± SD test values were lower in brachycephalic vs. nonbrachycephalic dogs (except for SMT; 7.4 ± 2.0 mm/5 seconds vs 7.3 ± 2.4 mm/5 seconds), with statistically significant differences noted for CTS (1.8 ± 0.5 cm vs 3.4 ± 0.8 cm), PRTT (37.2 ± 4.0 mm/15 seconds vs 41.1 ± 5.5 mm/15 seconds), STT-1 (20.1 ± 3.4 mm/min vs 23.3 ± 5.7 mm/min), STT-2 (13.0 ± 3.4 mm/min vs 16.9 ± 3.9 mm/min), and NL-STT1 (23.2 ± 3.6 mm/min vs 27.1 ± 5.4 mm/min), and nonsignificant differences for EAPPTT (16.6 ± 2.7 mm/15 seconds vs 17.5 ± 2.9 mm/15 seconds). Nasolacrimal stimulation increased STT-1 values by 18% on average. Correlations among tests were generally weak to moderate (r < .70) except for a strong correlation between STT-1 and NL-STT1 (r = .83, P < .001). Test reliability was good although test-retest repeatability was generally poor to moderate, as depicted by low intraclass correlation coefficients (ICC ≤ 0.75) and wide 95% limits of agreement, except for CTS (ICC = 0.91). CONCLUSIONS: Corneal sensitivity and aqueous tear secretion are lower in brachycephalic dogs. A comprehensive assessment of the ocular surface requires the combination of several diagnostic tests. The nasolacrimal reflex may provide a useful diagnostic and therapeutic tool in dogs.

The role of Xist-mediated Polycomb recruitment in the initiation of X-chromosome inactivation.

Xist RNA has been established as the master regulator of X-chromosome inactivation (XCI) in female eutherian mammals, but its mechanism of action remains unclear. By creating novel Xist-inducible mutants at the endogenous locus in male mouse embryonic stem (ES) cells, we dissect the role of the conserved A-B-C-F repeats in the initiation of XCI. We find that transcriptional silencing can be largely uncoupled from Polycomb repressive complex 1 and complex 2 (PRC1/2) recruitment, which requires B and C repeats. Xist ΔB+C RNA specifically loses interaction with PCGF3/5 subunits of PRC1, while binding of other Xist partners is largely unaffected. However, a slight relaxation of transcriptional silencing in Xist ΔB+C indicates a role for PRC1/2 proteins in early stabilization of gene repression. Distinct modules within the Xist RNA are therefore involved in the convergence of independent chromatin modification and gene repression pathways. In this context, Polycomb recruitment seems to be of moderate relevance in the initiation of silencing.

Conjunctival bacterial flora and antimicrobial susceptibility of captive and free-living sea turtles in Brazil.

PURPOSE: To describe the aerobic conjunctival bacterial flora of 3 especies of free-living and under human care sea turtles and determine its antimicrobial susceptibility in vitro. METHOD: Thirty-six sea turtles (72 eyes), juveniles and adults, 7 free-living Chelonia mydas and 8 Chelonia mydas, 4 Caretta caretta, 11 Eretmochelys imbricata, and 6 Lepidochelys olivacea under human care, were evaluated. Conjunctival cultures were collected for identification of aerobic bacteria and antimicrobial susceptibility testing for ciprofloxacin, chloramphenicol, gentamicin, neomycin, oxacillin, polymyxin B, tetracycline, and tobramycin using antibiotic disks. Bacterial strains showing no sensitivity to 4 or more antimicrobials were considered multiresistant to this panel. RESULTS: Bacterial growth was observed in 12/14 (85.71%) samples in the free-living sea turtles, and there was growth in 100% (58/58) of the samples from captive animals. There were 94 strains isolated and 15 species identified. There was a predominance of Gram-positive bacteria in free-living Chelonia mydas, most of which were Bacillus and Staphylococcus. The most commonly isolated Gram-negative species were enterobacteria for free-living and under human care animals. The strains were predominantly sensitive to ciprofloxacin and tobramycin, and less sensitive to oxacillin or polymyxin B. Ten multiresistant strains were isolated. Yeast were identified in 13.89% (10/72) of the samples. CONCLUSIONS: These results, showing differences in the conjunctival bacterial flora of free-living and captive animals, may be helpful for diagnosis and treatment of ocular disorders in sea turtles.