Vertebrate centromeres in mitosis are functionally bipartite structures stabilized by cohesin.

Centromeres are scaffolds for the assembly of kinetochores that ensure chromosome segregation during cell division. How vertebrate centromeres obtain a three-dimensional structure to accomplish their primary function is unclear. Using super-resolution imaging, capture-C, and polymer modeling, we show that vertebrate centromeres are partitioned by condensins into two subdomains during mitosis. The bipartite structure is found in human, mouse, and chicken cells and is therefore a fundamental feature of vertebrate centromeres. Super-resolution imaging and electron tomography reveal that bipartite centromeres assemble bipartite kinetochores, with each subdomain binding a distinct microtubule bundle. Cohesin links the centromere subdomains, limiting their separation in response to spindle forces and avoiding merotelic kinetochore-spindle attachments. Lagging chromosomes during cancer cell divisions frequently have merotelic attachments in which the centromere subdomains are separated and bioriented. Our work reveals a fundamental aspect of vertebrate centromere biology with implications for understanding the mechanisms that guarantee faithful chromosome segregation.

Vertebrate centromere architecture: from chromatin threads to functional structures.

Centromeres are chromatin structures specialized in sister chromatid cohesion, kinetochore assembly, and microtubule attachment during chromosome segregation. The regional centromere of vertebrates consists of long regions of highly repetitive sequences occupied by the Histone H3 variant CENP-A, and which are flanked by pericentromeres. The three-dimensional organization of centromeric chromatin is paramount for its functionality and its ability to withstand spindle forces. Alongside CENP-A, key contributors to the folding of this structure include components of the Constitutive Centromere-Associated Network (CCAN), the protein CENP-B, and condensin and cohesin complexes. Despite its importance, the intricate architecture of the regional centromere of vertebrates remains largely unknown. Recent advancements in long-read sequencing, super-resolution and cryo-electron microscopy, and chromosome conformation capture techniques have significantly improved our understanding of this structure at various levels, from the linear arrangement of centromeric sequences and their epigenetic landscape to their higher-order compaction. In this review, we discuss the latest insights on centromere organization and place them in the context of recent findings describing a bipartite higher-order organization of the centromere.

Short-Finned Pilot Whale Strandings Associated with Pilot Whale Morbillivirus, Brazil.

Cetacean morbillivirus (CeMV) causes illness and death in cetaceans worldwide; the CeMV strains circulating in the Southern Hemisphere are poorly known. We detected a pilot whale CeMV strain in 3 short-finned pilot whales (Globicephala macrorhynchus) stranded in Brazil during July-October 2020. Our results confirm this virus circulates in this species.

Molecular Detection and Characterization of Mycoplasma spp. in Marine Mammals, Brazil.

Mycoplasma spp. are wall-less bacteria able to infect mammals and are classified as hemotropic (hemoplasma) and nonhemotropic. In aquatic mammals, hemoplasma have been reported in California sea lions (Zalophus californianus) and river dolphins (Inia spp.). We investigated Mycoplasma spp. in blood samples of West Indian manatees (Trichechus manatus), pinnipeds (5 species), and marine cetaceans (18 species) that stranded or were undergoing rehabilitation in Brazil during 2002-2022. We detected Mycoplasma in blood of 18/130 (14.8%) cetaceans and 3/18 (16.6%) pinnipeds. All tested manatees were PCR-negative for Mycoplasma. Our findings indicate that >2 different hemoplasma species are circulating in cetaceans. The sequences from pinnipeds were similar to previously described sequences. We also detected a nonhemotropic Mycoplasma in 2 Franciscana dolphins (Pontoporia blainvillei) that might be associated with microscopic lesions. Because certain hemoplasmas can cause disease and death in immunosuppressed mammals, the bacteria could have conservation implications for already endangered aquatic mammals.

Hemotropic Mycoplasma spp. in Aquatic Mammals, Amazon Basin, Brazil.

Hemotropic Mycoplasma spp. (hemoplasmas) are uncultivable bacteria that infect mammals, including humans. We detected a potentially novel hemoplasma species in blood samples from wild river dolphins in the Amazon River Basin, Brazil. Further investigation could determine pathogenicity and zoonotic potential of the detected hemoplasma.

Myeloid and histiocytic sarcomas in subantarctic fur seals Arctocephalus tropicalis, Brazil.

Histopathological and immunohistochemical methods were used to diagnose round cell tumors in 2 subantarctic fur seals Arctocephalus tropicalis with marked anemia. Although wild-born, both individuals were placed under human care while juveniles in a Brazilian aquarium. Both pinnipeds were PCR tested for herpesvirus, and 1 was infected with otariid gammaherpesvirus 5 (OtHV-5), previously described in a subantarctic fur seal stranded in Brazil. Although some gammaherpesviruses can cause sarcomas and other neoplasms, it was not possible to definitively associate OtHV-5 with the neoplasm. To our knowledge, these are the first neoplasm records in subantarctic fur seals.

Spindle checkpoint silencing at kinetochores with submaximal microtubule occupancy.

The spindle assembly checkpoint (SAC) ensures proper chromosome segregation by monitoring kinetochore-microtubule interactions. SAC proteins are shed from kinetochores once stable attachments are achieved. Human kinetochores consist of hundreds of SAC protein recruitment modules and bind up to 20 microtubules, raising the question of how the SAC responds to intermediate attachment states. We show that one protein module ('RZZS-MAD1-MAD2') of the SAC is removed from kinetochores at low microtubule occupancy and remains absent at higher occupancies, while another module ('BUB1-BUBR1') is retained at substantial levels irrespective of attachment states. These behaviours reflect different silencing mechanisms: while BUB1 displacement is almost fully dependent on MPS1 inactivation, MAD1 (also known as MAD1L1) displacement is not. Artificially tuning the affinity of kinetochores for microtubules further shows that ∼50% occupancy is sufficient to shed MAD2 and silence the SAC. Kinetochores thus respond as a single unit to shut down SAC signalling at submaximal occupancy states, but retain one SAC module. This may ensure continued SAC silencing on kinetochores with fluctuating occupancy states while maintaining the ability for fast SAC re-activation.

High Occurrence of Chelonid Alphaherpesvirus 5 (ChHV5) in Green Sea Turtles Chelonia mydas with and without Fibropapillomatosis in Feeding Areas of the São Paulo Coast, Brazil.

Chelonid alphaherpesvirus 5 (ChHV5) has been consistently associated with fibropapillomatosis (FP), a neoplastic disease that affects sea turtles globally. The DNA of ChHV5 has been detected in cutaneous and noncutaneous tissues (e.g., lung) of green sea turtles Chelonia mydas with (FP+) and without (FP-) clinical signs of FP, indicating a persistent ChHV5 infection. Previously published and custom primer pairs were used to amplify the fragments of ChHV5 unique long (UL) partial genes (UL30 and UL18) through end-point PCR from cutaneous tumors (n = 31), nontumored skin (n = 49), and lungs (n = 26) from FP+ (n = 31) and FP- (n = 18) green sea turtles. The DNA of ChHV5 was detected in cutaneous tumors (80.6%, 25/31), nontumored skin (74.2%, 23/31 FP+; 27.8%, 5/18 FP-), and lung samples (91.7%, 11/12 FP+; 100%, 14/14 FP-). The high occurrence of ChHV5 observed in lung samples from FP- individuals was unexpected (14/14), providing the first evidence of ChHV5 DNA presence in lungs of individuals without FP. Our results also revealed high ChHV5 occurrence among the tested cohort (93.9%, 46/49) and suggested that a large proportion (83.4%, 15/18) of FP- green sea turtles had subclinical ChHV5 infections. Hence, our findings support the hypothesis that ChHV5 requires one or more possibly environmental or immune-related co-factors to induce FP.

Malassezia japonica is part of the cutaneous microbiome of free-ranging golden-headed lion tamarins (Leontopithecus chrysomelas - Kuhl, 1820).

We investigated Malassezia spp. in external ear canal and haircoat of free-ranging golden-headed lion tamarins (Leontopithecus chrysomelas). A total of 199 animals were restrained, and 597 clinical samples were collected. After the amplification of the 26S ribosomal gene by polymerase chain reaction (PCR), the RFLP technique was performed. Two additional PCR protocols were performed in 10 randomly selected strains. Malassezia sp. was isolated in 38.2% (76/199) of the animals and 14.6% (87/597) of the samples; all strains were lipodependent. The 10 sequenced strains showed a high identity with Malassezia japonica, species described in man, but not in animals, so far.

Postmortem findings in Magellanic penguins (Spheniscus magellanicus) caught in a drift gillnet.

BACKGROUND: Penguin interaction with gillnets has been extensively reported in the Atlantic and Pacific Oceans, and is considered a major conservation threat. Among penguin species, Magellanic penguins (Spheniscus magellanicus) are currently considered of great concern, particularly in Brazil, where they are highly susceptible to gillnet bycatch. Nevertheless, information about drowning-associated microscopic findings in penguins is limited. RESULTS: We describe the anatomopathological findings of 20 Magellanic penguins that drowned after getting entangled in a drift gillnet while wintering along the Brazilian shelf and washed ashore still enmeshed in Santa Catarina, Brazil. All 20 birds (19 juveniles and 1 adult; 18 females and 2 males) were in good body condition. Major gross findings were abrasion, bruising, and local erythema and edema of the wings, multiorgan congestion, jugular vein engorgement, pulmonary edema and hemorrhage, splenomegaly and hepatomegaly, fluid in the trachea, serous bloody fluid in the lungs, gastrointestinal parasites (nematodes, cestodes and trematodes), and debris in the stomach. The most common histopathological findings were cerebral and pulmonary congestion, pulmonary edema, splenic histiocytosis, lymphoid splenic hyperplasia, acute splenitis, extramedullary hepatic hematopoiesis, and parasitic enteritis. Although unspecific, the observed multiorgan congestion and pulmonary edema are consistent with previous reports of drowning in birds and may be indicative of this process. CONCLUSIONS: Drowning may be a challenging diagnosis (e.g., carcass decomposition, predation), but must be considered as a differential in all beach-cast seabird postmortem examinations. To the authors' knowledge this is the largest anatomopathological study based on microscopic examination in drowned penguins.

The Pathology of Cetacean Morbillivirus Infection and Comorbidities in Guiana Dolphins During an Unusual Mortality Event (Brazil, 2017-2018).

Cetacean morbillivirus (CeMV; Paramyxoviridae) is the most significant pathogen of cetaceans worldwide. The novel "multi-host" Guiana dolphin (Sotalia guianensis; GD)-CeMV strain is reported in South American waters and infects Guiana dolphins and southern right whales (Eubalaena australis). This study aimed to describe the pathologic findings, GD-CeMV viral antigen distribution and detection by RT-PCR (reverse transcriptase polymerase chain reaction), and infectious comorbidities in 29 Guiana dolphins that succumbed during an unusual mass-mortality event in Rio de Janeiro state, Brazil, between November 2017 and March 2018. The main gross findings were lack of ingesta, pulmonary edema, ascites, icterus, hepatic lipidosis, multicentric lymphadenomegaly, as well as pneumonia, polyserositis, and multiorgan vasculitis caused by Halocercus brasiliensis. Microscopically, the primary lesions were bronchointerstitial pneumonia and multicentric lymphoid depletion. The severity and extent of the lesions paralleled the distribution and intensity of morbilliviral antigen. For the first time in cetaceans, morbilliviral antigen was detected in salivary gland, optic nerve, heart, diaphragm, parietal and visceral epithelium of glomeruli, vulva, and thyroid gland. Viral antigen within circulating leukocytes suggested this as a mechanism of dissemination within the host. Comorbidities included disseminated toxoplasmosis, mycosis, ciliated protozoosis, and bacterial disease including brucellosis. These results provide strong evidence for GD-CeMV as the main cause of this unusual mass-mortality event.

Toxoplasmosis and Sarcocystis spp. infection in wild pinnipeds of the Brazilian coast.

The protozoans Toxoplasma gondii and Sarcocystis spp. (Sarcocystidae: Apicomplexa) affect a wide variety of vertebrates. Both have been reported to infect pinnipeds, with impacts on health ranging from inapparent to fulminant disease and death. However, little is known regarding their infections and associated pathology in South American pinnipeds. We used histological techniques to survey for the presence of T. gondii and Sarcocystis spp. in 51 stranded pinnipeds from Brazil. Immunohistochemical and molecular assays were employed in those cases consistent with Sarcocystidae infection. T. gondii cysts were detected in the central nervous system and heart of a South American fur seal Arctocephalus australis, associated with meningoencephalitis, myocarditis and endocarditis, and confirmed by immunohistochemistry. Additionally, this animal presented Sarcocystis sp. cysts in brain and heart tissues. Four additional specimens-2 Subantarctic fur seals A. tropicalis, an Antarctic fur seal A. gazella and another South American fur seal-presented intrasarcoplasmic cysts compatible with Sarcocystis spp. in muscle samples. There was no inflammation associated with the Sarcocystis spp. tissue cysts and all cysts were negative for S. neurona immunohistochemistry. The B1 gene of T. gondii was amplified in the 5 pinnipeds infected by Sarcocystidae protozoans. To our knowledge, this is the first report of toxoplasmosis in wild South American pinnipeds and of Sarcocystis spp. in South American fur seals. Detection of terrestrial parasites in aquatic mammals could be an indicator of their presence in the marine environment.

Ingression Progression Complexes Control Extracellular Matrix Remodelling during Cytokinesis in Budding Yeast.

Eukaryotic cells must coordinate contraction of the actomyosin ring at the division site together with ingression of the plasma membrane and remodelling of the extracellular matrix (ECM) to support cytokinesis, but the underlying mechanisms are still poorly understood. In eukaryotes, glycosyltransferases that synthesise ECM polysaccharides are emerging as key factors during cytokinesis. The budding yeast chitin synthase Chs2 makes the primary septum, a special layer of the ECM, which is an essential process during cell division. Here we isolated a group of actomyosin ring components that form complexes together with Chs2 at the cleavage site at the end of the cell cycle, which we named 'ingression progression complexes' (IPCs). In addition to type II myosin, the IQGAP protein Iqg1 and Chs2, IPCs contain the F-BAR protein Hof1, and the cytokinesis regulators Inn1 and Cyk3. We describe the molecular mechanism by which chitin synthase is activated by direct association of the C2 domain of Inn1, and the transglutaminase-like domain of Cyk3, with the catalytic domain of Chs2. We used an experimental system to find a previously unanticipated role for the C-terminus of Inn1 in preventing the untimely activation of Chs2 at the cleavage site until Cyk3 releases the block on Chs2 activity during late mitosis. These findings support a model for the co-ordinated regulation of cell division in budding yeast, in which IPCs play a central role.

Fusariosis in a Captive South American Sea Lion (Otaria flavescens): A Case Report.

Superficial mycoses are commonly reported in captive pinnipeds, usually maintained in wet and warm environments, favorable to fungal growth. Most superficial mycoses in pinnipeds have been described as difficult to treat; however, the majority of the reports come from past decades. Cutaneous lesions associated with opportunistic Fusarium sp. infections have been previously recognized in this taxon. We described the clinical signs, associated lesions and diagnosis (thermography, imprint cytology, histopathology, culture, electron microscopy, PCR) of a fusariosis case by Fusarium sp. in the nails and skin of an adult male captive South American sea lion (Otaria flavescens) recently transferred from another zoological institution, and its successful long-term treatment with Ketoconazole PO (60 days) and Miconazole solution spray TO, followed by Itraconazole PO (30 days). Herein we provide a successful approach to the diagnosis and treatment of fusariosis.

Survey of Brucella spp. and Leptospira spp. antibodies in cetaceans and manatees of the Amazon basin and Atlantic Ocean, Brazil.

Aquatic mammals can act as sentinels of emerging and resurging pathogens in the environment. Brucella spp. and Leptospira spp. are 2 zoonotic pathogens relevant to aquatic mammals, and their detection can be used to assess pathogen exposure. In this study, serum from 84 individuals-63 cetaceans (families Iniidae, n = 37; Delphinidae, n = 22; and Kogiidae, n = 4) and 21 West Indian manatees Trichechus manatus-was tested by the Rose Bengal Test (RBT) and a commercial competitive enzyme-linked immunosorbent assay (c-ELISA) for detecting Brucella spp. antibodies, and the microscopic agglutination test (MAT) for screening Leptospira spp. exposure. Overall, 4.8% (3/63) of cetaceans were positive by RBT and 15.9% (10/63) by c-ELISA for Brucella spp. Serum from 8 c-ELISA positive cetaceans (with available serum) was further tested via serum agglutination test (SAT) and 1 individual was positive. c-ELISA was more sensitive than RBT. Exposure to Brucella spp. was found in 5 cetacean species: Clymene dolphin Stenella clymene, short-finned pilot whale Globicephala macrorhynchus, pygmy killer whale Feresa attenuata, melon-headed whale Peponocephala electra and Atlantic bottlenose dolphin Tursiops truncatus in the Atlantic Ocean, Brazil, expanding the range of known Brucella seropositive aquatic hosts. No evidence of Brucella spp. exposure was found in Iniidae and Kogiidae odontocetes and manatees. Antibodies against Leptospira spp. were not detected in cetaceans and sirenians by MAT. These results contribute to the evaluation of different Brucella spp. serological methods in cetaceans and manatees and highlight the epidemiology of zoonotic pathogens in aquatic mammals of the southwestern Atlantic Ocean and the Amazon basin.

Pulmonary and systemic fungal infections in an Atlantic spotted dolphin and a Bryde's whale, Brazil.

We report the gross and microscopic findings and molecular identification of 2 cases of hyphate fungal infection in cetaceans from Brazil. The first case involved an adult male Atlantic spotted dolphin Stenella frontalis with localized pulmonary disease characterized by pyogranulomatous and necrotizing bronchopneumonia with intralesional hyphae. The second case involved an adult male Bryde's whale Balaenoptera edeni with orchitis, periorchitis, mesenteric lymphadenitis and pyogranulomatous bronchopneumonia with intralesional hyphae. PCR analysis from the dolphin's lung yielded Aspergillus fumigatus, and the fungus from the whale's mesenteric lymph node showed the greatest identity to Nanniziopsis obscura and Stagonosporopsis cucurbitacearum These cases represent the first reports of pulmonary aspergillosis by A. fumigatus in an Atlantic spotted dolphin and systemic mycosis by a possibly novel Onygenales in marine mammals.

Molecular identification and microscopic characterization of poxvirus in a Guiana dolphin and a common bottlenose dolphin, Brazil.

The poxviruses identified in cetaceans are associated with characteristic tattoo or ring skin lesions. However, little is known regarding the prevalence and progression of these lesions and the molecular characterization of cetacean poxviruses in the Southern Hemisphere. This manuscript describes the progression of poxvirus-like skin lesions in 5 free-ranging Guiana dolphins Sotalia guianensis. Additionally, 151 skin samples from 113 free-ranging cetaceans from Brazil, including 4 animals with tattoo skin lesions, were selected for poxvirus testing. Poxviral DNA polymerase gene PCR amplification was used to detect the virus in ß-actin-positive samples (145/151). DNA topoisomerase I gene PCR was then used in Cetaceanpoxvirus (CePV)-positive cases (n = 2), which were further evaluated by histopathology and electron microscopy. Based on photo-identification, adult Guiana dolphins presented regressing or healed poxvirus-like lesions (2/2), while juveniles presented persistent (2/3) or healed and progressive lesions (1/3). CePV DNA was amplified in a common bottlenose dolphin Tursiops truncatus and in a Guiana dolphin. Intracytoplasmic inclusion bodies and viral particles consistent with poxvirus were identified by histology and electron microscopy, respectively. CePV-specific amino acid motifs were identified through phylogenetic analysis. Our findings corroborate previous studies that suggest the placement of poxviruses from cetaceans within the novel CePV genus. This is the first molecular identification of poxvirus in South American odontocetes.

Imageability ratings across languages.

Imageability is a psycholinguistic variable that indicates how well a word gives rise to a mental image or sensory experience. Imageability ratings are used extensively in psycholinguistic, neuropsychological, and aphasiological studies. However, little formal knowledge exists about whether and how these ratings are associated between and within languages. Fifteen imageability databases were cross-correlated using nonparametric statistics. Some of these corresponded to unpublished data collected within a European research network-the Collaboration of Aphasia Trialists (COST IS1208). All but four of the correlations were significant. The average strength of the correlations (rho = .68) and the variance explained (R 2 = 46%) were moderate. This implies that factors other than imageability may explain 54% of the results. Imageability ratings often correlate across languages. Different possibly interacting factors may explain the moderate strength and variance explained in the correlations: (1) linguistic and cultural factors; (2) intrinsic differences between the databases; (3) range effects; (4) small numbers of words in each database, equivalent words, and participants; and (5) mean age of the participants. The results suggest that imageability ratings may be used cross-linguistically. However, further understanding of the factors explaining the variance in the correlations will be needed before research and practical recommendations can be made.

Lacaziosis-like disease in Tursiops truncatus from Brazil: a histopathological and immunohistochemical approach.

Cetacean lacaziosis-like disease or lobomycosis-like disease (LLD) is a chronic skin condition caused by a non-cultivable yeast of the order Onygenales, which also includes Lacazia loboi, as well as Paracoccidioides brasiliensis and P. lutzii, respectively responsible for lacaziosis and paracoccidioidomycosis in humans. Complete identification and phylogenetic classification of the LLD etiological agent still needs to be elucidated, but preliminary phylogenetic analyses have shown a closer relationship of the LLD agent to Paracoccidioides spp. than to L. loboi. Cases of LLD in South American cetaceans based on photographic identification have been reported; however, to date, only 3 histologically confirmed cases of LLD have been described. We evaluated multiple tissue samples from 4 Tursiops truncatus stranded in the states of Santa Catarina (n = 3) and Rio Grande do Sul (n = 1), southern Brazil. Macroscopically, all animals presented lesions consistent with LLD. Hematoxylin-eosin, periodic acid-Schiff, Grocott's methenamine silver, and Mayer's mucicarmin stains were used for histological evaluation. Microscopically, numerous refractile yeasts (4-9 µm in diameter) were observed in skin samples (4/4), and for the first time in dolphins, also in a skeletal muscle abscess (1/4). Immunohistochemistry using anti-P. brasiliensis glycoprotein gp43 as a primary antibody, which is known to cross-react with L. loboi and the LLD agent, was performed and results were positive in all 4 cases. We describe 3 new cases of LLD in cetaceans based on histopathology and immunohistochemistry. This is the first report of LLD in the muscle of cetaceans.