Bartonella spp. and Typhus Group Rickettsiae among Persons Experiencing Homelessness, São Paulo, Brazil.

Persons experiencing homelessness in São Paulo, Brazil, were seropositive for Bartonella spp. (79/109, 72.5%) and typhus group rickettsiae (40/109, 36.7%). Bartonella quintana DNA was detected in 17.1% (14/82) body louse pools and 0.9% (1/114) blood samples. Clinicians should consider vectorborne agents as potential causes of febrile syndromes in this population.

RNA sequencing identifies transcriptional changes in the rabbit larynx in response to low humidity challenge.

BACKGROUND: Voice disorders are a worldwide problem impacting human health, particularly for occupational voice users. Avoidance of surface dehydration is commonly prescribed as a protective factor against the development of dysphonia. The available literature inconclusively supports this practice and a biological mechanism for how surface dehydration of the laryngeal tissue affects voice has not been described. In this study, we used an in vivo male New Zealand white rabbit model to elucidate biological changes based on gene expression within the vocal folds from surface dehydration. Surface dehydration was induced by exposure to low humidity air (18.6% + 4.3%) for 8 h. Exposure to moderate humidity (43.0% + 4.3%) served as the control condition. Ilumina-based RNA sequencing was performed and used for transcriptome analysis with validation by RT-qPCR. RESULTS: There were 103 statistically significant differentially expressed genes identified through Cuffdiff with 61 genes meeting significance by both false discovery rate and fold change. Functional annotation enrichment and predicted protein interaction mapping showed enrichment of various loci, including cellular stress and inflammatory response, ciliary function, and keratinocyte development. Eight genes were selected for RT-qPCR validation. Matrix metalloproteinase 12 (MMP12) and macrophage cationic peptide 1 (MCP1) were significantly upregulated and an epithelial chloride channel protein (ECCP) was significantly downregulated after surface dehydration by RNA-Seq and RT-qPCR. Suprabasin (SPBN) and zinc activated cationic channel (ZACN) were marginally, but non-significantly down- and upregulated as evidenced by RT-qPCR, respectively. CONCLUSIONS: The data together support the notion that surface dehydration induces physiological changes in the vocal folds and justifies targeted analysis to further explore the underlying biology of compensatory fluid/ion flux and inflammatory mediators in response to airway surface dehydration.

Induction of acute graft vs. host disease in lymphopenic mice.

Allogeneic hematopoietic stem cell transplantation (HSCT) is a potentially life-saving treatment for refractory/relapsing hematological malignancies, blood disorders or autoimmune diseases. However, approximately 40-50% of patients undergoing allogeneic HSCT will develop a multi-organ, inflammatory disorder called acute graft vs. host disease (aGVHD). Experimental and clinical studies suggest that intestinal injury due to toxic, pre-transplant conditioning protocols (e.g. lethal irradiation and/or chemotherapy) may play a major role in the development of aGVHD. However, recent studies from our laboratory suggest that this may not be the case. The objective of this study was to quantify and compare the onset and severity of aGVHD induced by the adoptive transfer of allogeneic T cells into untreated lymphopenic mice. Four million allogeneic or syngeneic CD4+CD62L+CD25- T cells were transferred (i.p.) into NK cell-depleted RAG1-/- mice or RAG2-/-IL2rγ-/-double knock-out (DKO) mice and assessed daily for signs of aGVHD. We found that adoptive transfer of allogeneic but not syngeneic T cells into NK cell-depleted RAG1-/- or DKO mice induced many of the clinical and histological features of aGVHD including weight loss, inflammatory cytokine production and tissue inflammation. In addition, adoptive transfer of allogeneic T cells into each recipient induced severe anemia as well as dramatic reductions in bone marrow and spleen cellularity. Taken together, we conclude that allogeneic CD4+ T cells are both necessary and sufficient to induce aGVHD in lymphopenic recipients in the absence of toxic, pre-transplant conditioning.

Occurrence and identification of hemotropic mycoplasmas (Hemoplasmas) in free ranging and laboratory rats (Rattus norvegicus) from two Brazilian zoos.

BACKGROUND: Hemotropic mycoplasmas (hemoplasmas), bacteria belonging to the class Mollicutes, are obligatory red blood cell pathogens of a variety of animal species. They may cause acute anemia that is life-threatening or chronic disease that is clinically silent, but may interfere with results of experimental studies when using infected animals. Since these bacteria cannot be cultivated, molecular techniques are the gold standard for diagnosing an infection, investigating its prevalence, and describing new species. Mycoplasma coccoides and M. haemomuris are the most commonly recognized hemoplasmas in the blood of wild and laboratory rodents. Neither the epidemiology nor clinical and molecular characterization of hemoplasma infection in free-ranging rodents in Brazil has been previously reported. The aims of this study were to investigate the occurrence of hemoplasmas in free-ranging rats (Rattus norvegicus) captured in the Passeio Público and Curitiba Zoo and compare hematologic parameters of infected and non-infected animals. RESULTS: Anti-coagulated blood samples collected from 43 free-ranging and 20 nursery rats were included in the study. Overall 63.5% were positive using SYBR® Green quantitative PCR (qPCR) of the 16S rRNA gene to screen for hemoplasma infection (72% among free-ranging rats; 45% among laboratory-raised rats). Sequencing of the qPCR products showed that all but one sample had >98% identity to M. haemomuris. Phylogenetic analysis based on a fragment of approximately 1300 bp of the 16S rRNA gene showed 99% identity to a new hemoplasma from European rats and 98% identity to a hemotropic mycoplasma described infecting a European harvest mouse (Micromys minutus). No statistically significant changes in hematologic parameters between infected and non-infected rats were found, confirming the low pathogenicity and/or silent characteristics of the infection. CONCLUSIONS: Our findings suggest that hemoplasmas are likely endemic in rodent species in this region. The epidemiology, especially as it relates to the mode of transmission, needs to be further investigated as well as the possibility that other animal species, including humans, might become infected.

One Health approach to Coxiella burnetii in Brazilian indigenous communities.

Indigenous health has posted complex challenges worldwide, particularly due to historical economic, territorial, social and environmental processes, which may lead to emergence and reemergence of pathogens. In addition to few Coxiella burnetii serosurveys in vulnerable populations, especially in developing tropical countries, no comprehensive One Health approach has focused on human-animal infection along with potential environmental determinants. Accordingly, this study aimed to assess the seroprevalence of anti-C. burnetii antibodies in indigenous populations and their dogs from 10 indigenous communities distributed in southern and southeastern Brazil, along with the correspondent healthcare professionals. In overall, 8/893 (0.90%; 95% CI 0.45-1.76) indigenous and 1/406 (0.25%) dog samples were seropositive, with 7/343 (2.04%) individuals the 1/144 (0.69%) dog from the Ocoy community, located in the city of São Miguel do Iguaçu, bordering Argentina at south, and far 10 km at west from Paraguay. All 84 healthcare professionals tested seronegative.

A review on microRNA detection and expression studies in dogs.

MicroRNAs (miRNAs) are small non-coding RNAs that function by post-transcriptional regulation of gene expression. Their stability and abundance in tissue and body fluids makes them promising potential tools for both the diagnosis and prognosis of diseases and attractive therapeutic targets in humans and dogs. Studies of miRNA expression in normal and disease processes in dogs are scarce compared to studies published on miRNA expression in human disease. In this literature review, we identified 461 peer-reviewed papers from database searches using the terms "canine," "dog," "miRNA," and "microRNA"; we screened 244 for inclusion criteria and then included a total of 148 original research peer-reviewed publications relating to specific miRNA expression in canine samples. We found an overlap of miRNA expression changes between the four groups evaluated (normal processes, non-infectious and non-inflammatory conditions, infectious and/or inflammatory conditions, and neoplasia) in 39 miRNAs, 83 miRNAs in three of the four groups, 110 miRNAs in two of the three groups, where 158 miRNAs have only been reported in one of the groups. Additionally, the mechanism of action of these overlapping miRNAs varies depending on the disease process, elucidating a need for characterization of the mechanism of action of each miRNA in each disease process being evaluated. Herein we also draw attention to the lack of standardization of miRNA evaluation, consistency within a single evaluation method, and the need for standardized methods for a direct comparison.

Comparative proteomic changes in rabbit vocal folds undergoing systemic dehydration and systemic rehydration.

BACKGROUND: A considerable body of clinical evidence suggests that systemic dehydration can negatively affect voice production, leading to the common recommendation to rehydrate. Evidence for the corrective benefits of rehydration, however, is limited with mixed conclusions, and biological data on the underlying tissue changes with rehydration is lacking. In this study, we used a rabbit model (n = 24) of acute (5 days) water restriction-induced systemic dehydration with subsequent rehydration (3 days) to explore the protein-level changes underlying the molecular transition from euhydration to dehydration and following rehydration using LC-MS/MS protein quantification in the vocal folds. We show that 5-day water restriction led to an average 4.3% decrease in body weight with relative increases in anion gap, Cl-, creatinine, Na+, and relative decreases in BUN, iCa2+, K+, and tCO2 compared to control (euhydrated) animals. A total of 309 differentially regulated (p < 0.05) proteins were identified between the Control and Dehydration groups. We observed a noteworthy similarity between the Dehydration and Rehydration groups, both well differentiated from the Control group, highlighting the distinct timelines of resolution of the clinical symptoms of systemic dehydration and the underlying molecular changes. SIGNIFICANCE: Voice disorders are a ubiquitous problem with considerable economic and psychological impact. Maintenance of proper hydration is commonly prescribed as a general vocal hygiene practice. There is evidence that dehydration negatively impacts phonation, but our understanding of the state of vocal folds in the context of systemic dehydration are limited, particular from a molecular perspective. Further, ours is a novel molecular study of the short-term impact of rehydration on the tissue. Given the relatively minimal difference in vocal fold proteomic profiles between the Dehydration and Rehydration groups, our data demonstrate a complex physiological response to acute systemic dehydration, and highlight the importance of considering persistent underlying molecular pathology despite the rapid resolution of clinical measures. This study sets a foundation for future research to confirm the nature of potential beneficial outcomes of clinical recommendations related to hydration.

Influence of Housing Temperature and Genetic Diversity on Allogeneic T Cell-Induced Tissue Damage in Mice.

The objective of this study was to determine how housing temperature and genetic diversity affect the onset and severity of allogeneic T cell-induced tissue damage in mice subjected to reduced intensity conditioning (RIC). We found that adoptive transfer of allogeneic CD4+ T cells from inbred donors into sub-lethally irradiated inbred recipients (I→I) housed at standard housing temperatures (ST; 22-24 °C) induced extensive BM and spleen damage in the absence of injury to any other tissue. Although engraftment of T cells in RIC-treated mice housed at their thermo-neutral temperature (TNT; 30-32 °C) also developed similar BM and spleen damage, their survival was markedly and significantly increased when compared to their ST counterparts. In contrast, the adoptive transfer of allogeneic T cells into RIC-treated outbred CD1 recipients failed to induce disease in any tissue at ST or TNT. The lack of tissue damage was not due to defects in donor T cell trafficking to BM or spleen but was associated with the presence of large numbers of B cells and myeloid cells within these tissues that are known to contain immunosuppressive regulatory B cells and myeloid-derived suppressor cells. These data demonstrate, for the first time, that housing temperature affects the survival of RIC-treated I→I mice and that RIC-conditioned outbred mice are resistant to allogeneic T cell-induced BM and spleen damage.

Molecular detection of feline hemoplasmas and retroviruses in free-roaming and shelter cats within a university campus.

Objectives: The aim of the present study was to assess the frequency of hemoplasma, feline immunodeficiency virus (FIV) and feline leukemia virus (FeLV) infections in cats living in an on-campus shelter and free-roaming cats within a university campus in Brazil. Methods: Blood samples were tested using quantitative PCR for hemoplasma, FIV and FeLV. Positive hemoplasma samples were sequenced. Associations between hemoplasma detection and living situation, sex, flea and/or tick parasitism, and coinfection with FIV and FeLV, were assessed using Fisher's exact test and the respective odds ratios were calculated. Results: Overall, 6/45 (13.3%) cats tested positive: four (8.9%) were infected with 'Candidatus Mycoplasma haemominutum' and two (4.4%) with Mycoplasma haemofelis. All positive samples were from free-roaming cats (6/15; 40.0%) and had statistically significantly lower packed cell volumes (P = 0.037). Although 5/23 (21.7%) males and 1/22 (4.6%) females were positive, no statistically significant association between sex and hemoplasma infection was found (P = 0.19). Viral quantitative PCR (qPCR) was performed on 43/45 samples, among which 2/43 (4.7%) were positive for FIV and none for FeLV. Only one cat (2.3%) was coinfected with hemoplasma and FIV (P = 0.26). In addition, 4/6 (66.7%) cats that tested positive for hemoplasmas were infested by fleas (P = 0.0014) and/or ticks (P = 0.25). Conclusions and relevance: These results show that even if the free-roaming cat population is clinically healthy and has adequate access to food, it may present flea infestation and hemoplasma infection with lower packed cell volume values.

Seroprevalence and associated risk factors of strongyloidiasis in indigenous communities and healthcare professionals from Brazil.

Strongyloides stercoralis, a pathogenic roundworm, is considered endemic in several tropical and subtropical areas worldwide. Indigenous populations have the highest soil-transmitted helminthiases-related mortality rates, but the prevalence and risk factors associated with S. stercoralis in Brazilian indigenous populations have not been established. Thus, the present study aimed to assess the seroprevalence and associated risk factors for S. stercoralis in indigenous communities and the healthcare professionals serving them in Brazil. Indigenous populations living in nine communities and healthcare professionals were tested for anti- S. stercoralis antibodies by ELISA. A questionnaire was used to assess socio-epidemiological information. Associated risk factors for seropositivity were tested by chi-square or Fisher's exact tests, using univariate analyses and multivariate logistic regression. Overall, 174/463 (37.6%; CI 95%: 33.3-42.1) indigenous persons and 77/147 (52.4%; 95% CI: 44.3-60.3) healthcare professionals were seropositive for anti- S. stercoralis antibodies. Seropositivity among the two groups was statistically significant (p = 0.0016; OR = 0.547; 95% CI: 0.376-0.796) and revealed that healthcare professionals were 1.83 times more likely to be seropositive. The multivariate analysis showed that being male or being adult were also risk factors, while having a septic tank as a sanitary facility represented a protective factor for S. stercoralis exposure in indigenous persons. None of the variables evaluated were associated with S. stercoralis exposure in the professional group. The study herein has reported a high seroprevalence to Strongyloides stercoralis in indigenous communities of Brazil and healthcare professionals, warning for potential public health concerns of strongyloidiasis in such populations.

Risk factors for toxocariasis during incarceration: the One Health intervention approach.

Despite potential exposure to soil-transmitted helminths, especially when stray dogs and cats are present, toxocariasis in inmate populations remains to be established. Accordingly, the present study assessed the seroprevalence and associated risk factors of toxocariasis at the Women's State Penitentiary of Parana, Brazil. A total of 234/370 (63.2%; 95% CI 58.2-68.0) women inmates and 28/87 (32.2%; 95% CI 23.3-42.6) correctional officers were seropositive for anti-Toxocara spp. IgG by ELISA, with inmates 2.62-fold more likely positive (p = 0.00000026). The univariate model has identified that non-white (OR = 1.58, p = 0.047) and older than 39 years (OR = 1.28, p = 0.032) inmates were associated with mild but significant odds for seropositivity. Elementary or higher educational level was considered a protective factor for seropositivity. The presence of Toxocara spp. eggs was observed in 10/15 (66.7%) collected soil samples by centrifuge-flotation in Zinc Sulfate, and molecular analysis by PCR identified only Toxocara cati in these eggs. An intervention program was established with regular trap-neuter-release, with gradual removal for adoption (donation campaigns), treatment, and euthanasia when necessary (particularly due to advanced sporotrichosis). In addition, an educational awareness agenda was proposed, aiming to reduce soil contamination and accidental intake by the incarcerated population. A total of 40 feral cats were trapped, 20 males and 20 females, mostly adults. After trapping, 36 cats were neutered, treated, and microchipped in the Veterinary Teaching Hospital (VTH) at the Federal University of Paraná. Five trapped feral cats were euthanized, four diagnosed with advanced sporotrichosis, and one already neutered cat (not herein) with complications due to feline immunodeficiency virus (FIV). Female inmates presented higher seroprevalence for Toxocara spp. antibodies when compared to correctional officers, significantly associated with age, self-declared ethnicity (non-white), and lack of formal education. Despite the non-natural scenario of a state penitentiary, the One Health approach of Toxocara spp. has highlighted the interdisciplinary nature of the study and its relevance in understanding the complex interactions between human, animal, and environmental factors, particularly impacting female inmates. Further studies should establish the rate of inmate infection over time while deprived of liberty.

Toxoplasma gondii exposure in Brazilian indigenous populations, their dogs, environment, and healthcare professionals.

Although Toxoplasma gondii exposure has been reported in indigenous populations worldwide, a One Health approach has not been applied to date. This study concurrently assessed T. gondii exposure in indigenous populations, and their dogs, environment, and indigenous or non-indigenous healthcare professionals (HPs). Human and dog serum samples from 9 indigenous communities in Brazil were assessed by indirect immunofluorescence antibody test for anti-T. gondii antibodies. Soil samples (30 per community) were processed with PCR to amplify T. gondii DNA. Associated risk factors and seroprevalence were analyzed using logistic regression models. Human seropositivity and type of water source were assessed by generalized linear mixed model (GLMM) with binomial error distribution, and game meat consumption with chi-squared test. Overall, 225/463 (49%) indigenous persons were seropositive for anti-T. gondii antibodies. Of all the HPs, 67/168 (40%) were positive, and included 54/147 (37%) positive non-indigenous HPs. Indigenous persons more likely to be seropositive compared with non-indigenous HPs (OR: 1.63; 95% CI: 1.11-2.39). A total of 97/253 (38%) dogs were seropositive and highly associated with seropositive owners (p < 0.001). Based on univariate analysis for indigenous individuals, state location of community (p < 0.001), ethnicity (p < 0.001), consumption of game meat (p < 0.001), type of water source (p < 0.001), and educational level (p = 0.026) were associated with seropositivity. Logistic regression showed that indigenous seropositivity was associated with eating game meat (p = 0.002), drinking water from rivers (p < 0.001), and inversely proportional to the educational level. According to univariate analysis for non-indigenous HP, age (p = 0.005), frequency of visits to the indigenous populations (p < 0.001), consumption of water at the indigenous communities (p < 0.001), and ingestion of raw meat (p = 0.023) were associated with T. gondii seropositivity. Logistic regression revealed living outdoors (p = 0.042), habit of hunting (p = 0.008), and drinking river water (p = 0.007) as risk factors associated to seropositivity in dogs. In addition, indigenous communities lacking water treatment had higher seroprevalence for all groups including indigenous persons (GLMM; z = -7.153; p < 0.001), their dogs (GLMM; z = -2.405; p = 0.0162), and all HPs (GLMM; z = -2.420; p = 0.0155). Human seropositivity was associated with that of their dogs (p < 0.001). A single soil sample, out of 270 (0.37%), was positive for T. gondii by PCR. Our results indicate water source is a risk for human and dog toxoplasmosis in indigenous communities; both share similar exposure. Moreover, quality water access was shown to be crucial to prevent toxoplasmosis in both total and non-indigenous HPs who work in these indigenous communities.

One health approach to toxocariasis in Brazilian indigenous populations, their dogs, and soil contamination.

Introduction: Although socioeconomic vulnerability and lifestyle factors may contribute to the transmission of Toxocara spp., no study has investigated indigenous populations in Brazil using the One Health approach. Methods: Accordingly, this study assessed anti-Toxocara spp. antibodies in Brazilian indigenous people and healthcare professionals by enzyme-linked immunosorbent assay. Presence of Toxocara spp. eggs (feces and hair) in dogs as definitive hosts and in soil samples of the indigenous communities were also recovered and molecularly investigated. Results: Overall, 342/463 (73.9%) indigenous individuals and 46/147 (31.3%) non-indigenous healthcare professionals were seropositive for Toxocara spp. In addition, T. canis eggs were retrieved from 9/194 (4.6%) dog fecal samples and 4/204 (2.0%) dog hair samples, mainly from the Paraná State communities (3/42; 7.1%). Soil contamination was observed only in the Paraná State communities (36/90; 40.0%), with the molecular detection of T. canis. River water consumption was also associated with indigenous seropositivity (Odds ratio, 11.4). Discussion: Indigenous individuals in Paraná State communities were 2.72-fold more likely to be seropositive than those in São Paulo State, likely due to a lack of sanitary infrastructure. In this scenario, a primarily soil-transmitted disease may also have become waterborne, with embryonated eggs probably spread to water supplies by rain. Full-time healthcare professionals in daily contact with indigenous communities were 9.2-fold more likely to be seropositive than professionals who visited sporadically, suggesting exposure to Toxocara spp. during their work and raising health concerns. In addition, the findings herein showed a significantly higher seroprevalence in indigenous people than in healthcare workers (χ2 = 85.5; p < 0.0001), likely due to overtime exposure to Toxocara spp. In conclusion, Brazilian indigenous communities are highly exposed to toxocariasis, with poor infrastructure and contact with contaminated river water as associated risk factors and a higher risk of infection in healthcare professionals working full-time in these communities.

Molecular detection and characterization of SARS-CoV-2 in cats and dogs of positive owners during the first COVID-19 wave in Brazil.

Despite previous reports of SARS-CoV-2 infection in dogs and cats worldwide, the type of swab sample used for its detection through RT-qPCR needs to be better compared and described. Accordingly, as part of a multicenter study in Brazil, the aim of the present study was to assess which rectal or oropharyngeal swabs would be more appropriate for detecting SARS-CoV-2 in cats and dogs, through viral load comparison. Pets of owners diagnosed with COVID-19 in the last 7 days were eligible. A total of 148 animals from four of the five Brazilian geographical regions were analyzed, among which 10/48 cats (20.83%) and 11/100 dogs (11.00%) were positive. The results suggested that oropharyngeal swabs should be considered for SARS-CoV-2 detection, particularly in cats, due to the higher cDNA viral load. Also, the genomic results showed similarities between SARS-CoV-2 animal variants and human variants that were circulating at the time of sampling, thus corroborating the existence of zooanthroponotic transmission. In conclusion, the present study highlighted the importance of SARS-CoV-2 monitoring among cats and dogs, as virus modification may indicate the possibility of mutations in animals and spillover back to owners. Thus, positive individuals should always self-isolate from their pets during COVID-19, to prevent trans-species transmission and mutation.

Hemophagocytic syndrome associated with large granular lymphoma in an adult dog.

Hemophagocytic syndrome (HPS) is a rare disorder characterized by dysregulation of the immune response resulting in uncontrolled activation of macrophages with exacerbated phagocytosis of host cells. In dogs, the criteria for diagnosis include the presence of pancytopenia or bicytopenia in the peripheral blood and >2% hemophagocytic macrophages in bone marrow aspirates. When HPS is associated with lymphoma, it is called lymphoma-associated hemophagocytic syndrome (LAHS). Here, we present a case of a 4 ½-year-old female spayed Old English Mastiff that presented with severe thrombocytopenia, mild anemia, mild to moderate leukopenia, and large granular lymphocytes (LGLs) in the peripheral blood. The patient had enlarged lymph nodes with many LGLs seen cytologically, leading to the interpretation of LGL lymphoma. Bone marrow displayed numerous LGLs that stained strongly for CD3 but did not show immunoreactivity to CD4 or CD8, and PCR for antigen receptor rearrangement analysis confirmed a clonal T-cell receptor gamma gene rearrangement. The presence of ~3.5% hemophagocytes present on the bone marrow evaluation raised concern for HPS and, more specifically, LAHS. HPS and LAHS are challenging to diagnose and require many criteria to be fulfilled before a definitive diagnosis can be made; the low number of cases in the literature makes this even more challenging in dogs. This case represents secondary LAHS due to LGL lymphoma in a dog.

Furosemide-induced systemic dehydration alters the proteome of rabbit vocal folds.

Whole-body dehydration (i.e., systemic dehydration) leads to vocal fold tissue dehydration. Furosemide, a common diuretic prescribed to treat hypertension and edema-associated conditions, induces systemic dehydration. Furosemide also causes voice changes in human speakers, making this method of systemic dehydration particularly interesting for vocal fold dehydration studies. Our objective was to obtain a comprehensive proteome of vocal folds following furosemide-induced systemic dehydration. New Zealand White rabbits were used as the animal model and randomly assigned to euhydrated (control) or furosemide-dehydrated groups. Systemic dehydration, induced by injectable furosemide, was verified by an average body weight loss of -5.5% and significant percentage changes in blood analytes in the dehydrated rabbits compared to controls. Vocal fold specimens, including mucosa and muscle, were processed for proteomic analysis using label-free quantitation LC-MS/MS. Over 1600 proteins were successfully identified across all vocal fold samples; and associated with a variety of cellular components and ubiquitous cell functions. Protein levels were compared between groups showing 32 proteins differentially regulated (p ≤ 0.05) in the dehydrated vocal folds. These are mainly involved with mitochondrial translation and metabolism. The downregulation of proteins involved in mitochondrial metabolism in the vocal folds suggests a mechanism to prevent oxidative stress associated with systemic dehydration. SIGNIFICANCE: Voice disorders affect different population demographics worldwide with one in 13 adults in the United States reporting voice problems annually. Vocal fold systemic hydration is clinically recognized for preventing and treating voice problems and depends on optimal body hydration primarily achieved by water intake. Herein, we use the rabbit as a translatable animal model, and furosemide as a translatable method of systemic dehydration, to reveal a comprehensive proteomic profile of vocal fold mucosa and muscle in response to systemic dehydration. The significant subset of proteins differentially regulated due to furosemide-induced dehydration offer novel insights into the molecular mechanisms of systemic dehydration in the vocal folds. These findings also deepen our understanding of changes to tissue biology after diuretic administration.

One Health Index (OHI) applied to Curitiba, the ninth-largest metropolitan area of Brazil, with concomitant assessment of animal, environmental, and human health indicators.

One Health has been defined as a comprehensive approach to human, animal, and environmental health, but unsuccessful in providing a specific index for city, state, or country assessment. Accordingly, the present study aimed to calculate the One Health Index (OHI) in 29 cities of the Curitiba metropolitan area, the ninth largest in Brazil. Animal and Environmental health indicators were obtained from the city secretary of environment. The social vulnerability index (SVI) was used as the overall human health indicator. The indicators were parameterized following a binary logic Principal component analysis (PCA) was applied in association with city population, per capita income, and distance from the capital Curitiba. Permutational multivariate analysis of variance (PERMANOVA) was applied using the three first coefficients of the principal components obtained from Principal Component Analysis Plot (PCA) and exploring a pairwise comparison between the scenario ranges. A value of p less than 0.05 was considered significant. Overall, a higher OHI was associated with a higher city population and income, and shorter distance from the capital, and tendency of low-income cities to present lower OHI compared to higher-income cities. In conclusion, the OHI proposed herein portrays a holistic representation of a city's overall health. Moreover, animal issues should be considered part of local to global sustainability, considering OHI to calculate sustainability indexes.

Novel duplex RT-qPCR for animal rabies surveillance.

Rabies is a lethal zoonosis affecting mammals worldwide. Diagnosis of rabies follows international standard protocols, primarily relying on direct immunofluorescence (DI) followed by mouse inoculation test (MIT). WHO recommends molecular biology techniques such as RT-qPCR for replacing MIT to diagnose rabies in animal samples. Recently, a real-time PCR protocol that detects all rabies virus variants identified worldwide was validated. This assay is a pan-Lyssavirus TaqMan quantitative RT-PCR called LN34. A modified LN34 assay protocol was tested at the Paraná State Reference Laboratory (Lacen/PR) using animal samples previously tested by DI and MIT, the gold standard (GS). This method has been changed to a RT-qPCR duplex format to better fit the diagnostic routine. The new assay was called duplex LN34 and ß-actin RT-qPCR. All the 88 samples evaluated using the GS test, modified pan-Lyssavirus TaqMan RT-qPCR and duplex LN34 and ß-actin RT-qPCR showed 100% agreement with each other. This novel duplex RT-qPCR protocol has shown adequate diagnostic performance and may be used in research and surveillance purposes, replacing the standard MIT and ending mice use for rabies diagnosis.

Serological survey of anti-Leptospira spp. antibodies in individuals with animal hoarding disorder and their dogs in a major city of Southern Brazil.

BACKGROUND: Individuals with hoarding disorder (HD) presented a persistent difficulty in detaching from objects and/or animals. Unhealthy conditions, frequently found in cases of animal HD (AHD), may favour environmental contamination and the spread of zoonotic pathogens. Despite that, only one study of zoonotic diseases in individuals with AHD and their companion animals has been conducted to date. OBJECTIVES: This study aimed to assess the seroprevalence of anti-Leptospira spp. antibodies in individuals with AHD and their dogs in a major city of Southern Brazil. METHODS: Blood samples were obtained from 264 dogs (21 households) and 19 individuals with AHD (11 households) and tested by microscopic agglutination test. RESULTS: All human samples were seronegative. Seropositivity was found in 16/264 (6.1%; CI 95% 3.3-9.6%) dogs from 11/21 (52.38%) households, with titres ranging from 100 up to 400, and Copenhageni (10/16; 62.5%) was the most frequent serovar. Surprisingly, seropositivity of hoarded dogs found herein was among the lowest reportedly observed in other dog populations of Brazil. Two epidemiological variables were significantly associated with seropositivity in dogs: the presence of cat hoarding (p = 0.004) and the report of flood occurrence in the household (p = 0.031). CONCLUSIONS: No individuals with AHD were seropositive, and besides the lower seroprevalence of dogs, they probably had contact with Leptospira spp. at some point in their life. Since dogs can be considered potential sentinels in leptospirosis, public health programs must become aware of the risk of leptospirosis cases in households of individuals with AHD and nearby communities.

The rate and role of pseudogenes of the Mycobacterium tuberculosis complex.

Whole-genome sequence analyses have significantly contributed to the understanding of virulence and evolution of the Mycobacterium tuberculosis complex (MTBC), the causative pathogens of tuberculosis. Most MTBC evolutionary studies are focused on single nucleotide polymorphisms and deletions, but rare studies have evaluated gene content, whereas none has comprehensively evaluated pseudogenes. Accordingly, we describe an extensive study focused on quantifying and predicting possible functions of MTBC and Mycobacterium canettii pseudogenes. Using NCBI's PGAP-detected pseudogenes, we analysed 25 837 pseudogenes from 158 MTBC and M. canetii strains and combined transcriptomics and proteomics of M. tuberculosis H37Rv to gain insights about pseudogenes' expression. Our results indicate significant variability concerning rate and conservancy of in silico predicted pseudogenes among different ecotypes and lineages of tuberculous mycobacteria and pseudogenization of important virulence factors and genes of the metabolism and antimicrobial resistance/tolerance. We show that in silico predicted pseudogenes contribute considerably to MTBC genetic diversity at the population level. Moreover, the transcription machinery of M. tuberculosis can fully transcribe most pseudogenes, indicating intact promoters and recent pseudogene evolutionary emergence. Proteomics of M. tuberculosis and close evaluation of mutational lesions driving pseudogenization suggest that few in silico predicted pseudogenes are likely capable of neofunctionalization, nonsense mutation reversal, or phase variation, contradicting the classical definition of pseudogenes. Such findings indicate that genome annotation should be accompanied by proteomics and protein function assays to improve its accuracy. While indels and insertion sequences are the main drivers of the observed mutational lesions in these species, population bottlenecks and genetic drift are likely the evolutionary processes acting on pseudogenes' emergence over time. Our findings unveil a new perspective on MTBC's evolution and genetic diversity.