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An unprecedented outbreak of rabies occurred in Rio Grande do Sul state (RS) from 2012 onward, resulting in thousands of bovine deaths, important economic losses, and posing risk to human health. This article describes a genetic analysis of 145 rabies viruses (RABV) recovered from herbivorous from RS between 2012 and 2017, based on partial sequence analysis of the nucleoprotein (N) gene. High nucleotide ( nt ) identity (95.5 to 100%) and amino acid ( aa ) similarity (96.7 to 100%) were observed among the analyzed sequences. These sequences displayed a high sequence nt identity/ aa similarity with bovine RABV sequences (96.4–97.9%; 98.1–100%, respectively) and vampire bat RABV sequences (96.3–97.5%; 97.8–99.5%). Phylogenetic analyzes based on the N sequence allowed for the segregation of viruses into two distinct clusters . Cluster 1 comprised RABV sequences covering the whole studied period, whereas cluster 2 grouped a lower number of viruses from 2013, 2014, 2015, to 2017. In some cases, viruses obtained from the same region within a short period of time grouped to distinct clusters or sub-clusters, indicating the co-circulation of distinct virus lineages in these outbreaks. The segregation into sub-clusters was also observed for viral sequences obtained from the same region at different times, indicating the involvement of distinct viruses. In summary, partial sequence analyses revealed a high conservation of N protein and the circulation of two lineages and different sublineages of RABV in the region. In addition, our results confirm the suitability of N gene to study the genetic relationships among RABV isolates.

Discussion section, 9th paragraph.

A large outbreak of hematophagous-bat-associated bovine rabies has been occurring in Rio Grande do Sul (RS), the southernmost Brazilian state, since 2011, with official estimates exceeding 50,000 cattle deaths. The present article describes a genetic characterization of rabies virus (RABV) recovered from 59 affected cattle and two sheep, from 56 herds in 16 municipalities (2012-2016). Molecular analysis was performed using the nucleotide (nt) and predicted amino acid (aa) sequences of RABV glycoprotein G (G). A high level of nt and aa sequence identity was observed among the examined G sequences, ranging from 98.4 to 100%, and from 97.3 to 100%, respectively. Likewise, high levels of nt and aa sequence identity were observed with bovine (nt, 99.8%; aa, 99.8%) and hematophagous bat (nt, 99.5%; aa, 99.4%) RABV sequences from GenBank, and lower levels were observed with carnivore RABV sequences (nt, 92.8%; aa, 88.1%). Some random mutations were observed in the analyzed sequences, and a few consistent mutations were observed in some sequences belonging to cluster 2, subcluster 2b. The clustering of the sequences was observed in a phylogenetic tree, where two distinct clusters were evident. Cluster 1 comprised RABV sequences covering the entire study period (2012 to 2016), but subclusters corresponding to different years could be identified, indicating virus evolution and/or introduction of new viruses into the population. In some cases, viruses from the same location obtained within a short period grouped into different subclusters, suggesting co-circulation of viruses of different origins. Subcluster segregation was also observed in sequences obtained in the same region during different periods, indicating the involvement of different viruses in the cases at different times. In summary, our results indicate that the outbreaks occurring in RS (2012 to 2016) probably involved RABV of different origins, in addition to a possible evolution of RABV isolates within this period.

Bats and dogs are the main reservoirs of rabies virus (RABV) in Latin America and are responsible for the maintenance of different cycles of infection. In the two neighbour and most southern Brazilian states of Rio Grande do Sul (RS) and Santa Catarina (SC), rabies in dogs has been successfully controlled for more than 30 years. However, rabies associated to the rural cycle remains endemic, with a significant, though oscillating—annual incidence of rabies in cattle. Despite the plethora of studies on genetic analyses of Brazilian RABV, isolates from southern Brazil have only scarcely been investigated. This work was performed to identify the genetic lineages of RABVs circulating in states of RS and SC. Fifty-nine RABV cattle isolates from RS and SC were selected and submitted to reverse transcription/polymerase chain reaction (RT-PCR) followed by sequencing of the nucleoprotein gene. In RS, the circulation of two sublineages (1A and 1B) of RABV was detected, both with characteristics of lineages usually detected in vampire bats (Desmodus rotundus). In SC, only one sublineage of RABV (1B) was detected. Nevertheless, the findings reported here are expected to contribute to the understanding of the biology of the virus in the region and its interactions with the natural host D. rotundus.

This study evaluated the effects of restricted nursing (RN) and the administration of a bovine appeasing substance (BAS) given 8 days before (d‐8) and on the day of weaning (d0) as a strategy to mitigate stress and optimize the post‐weaning growth performance of beef calves, with emphasis on behavior and response to vaccination. Forty male Nelore calves were randomly assigned to four treatments with ten replicates: control (CON)—topical administration of 5 mL of saline solution on d0; BAS‐8—topical administration of 5 mL of BAS on d‐8; BAS0—topical administration of 5 mL of BAS on d0; and RN—calves allowed to nurse only one hour/day from d‐8 to d0. Calves in the CON treatment group showed lower average daily gain compared to the other treatments ( p  < 0.05). There was a trend ( p  = 0.09) for BAS‐8 and BAS0 to result in lower temperament scores when calves entered the restraint chute. BAS0 calves had greater ( p  = 0.05) serum titer concentrations against rabies compared to the other treatments. The RN and BAS administration eight days before and on the day of weaning was effective in improving growth performance in beef calves.

In the intestinal lumen, excess of oxides and sulfates interfere with the absorption of minerals due to competition from the same absorption site. Amino acids-mineral complexed (AACM) is intended to minimize these problems, which might be absorbed by different absorption sites. Then, a study including Zinc (Zn), Manganese (Mn) and Copper (Cu) from different sources was carried out to evaluate the performance, blood parameters and reproductive organs development of Brown Laying Hens. A total of 800 Lohmann Brown Lite were fed, from one-day-old to 182-days-old, Zn, Mn and Cu from different sources. Measurements were made from 105 to 182-days-old. The laying hens were distributed according to a completely randomized design with 20 replicates and 20 birds per experimental unit. The treatments consisted of a diet supplemented with 70, 70 and 8 mg/kg of Zn, Mn and Cu; respectively, from inorganic sources (IM). The second treatment contained 40, 40 and 2.75 mg/kg of Zn, Mn and Cu, respectively from IM plus 30, 30 and 5.25 mg/kg of Zn, Mn and Cu; respectively, from AACM sources. Performance and reproductive organs development (oviduct and ovary weight), tibia weight, liver weight, egg output and body weight, and blood variables were evaluated. Data were compared by Student's t-test (P < 0.05). Laying hens fed AACM reached 35% of egg output two days earlier and presented heavier tibia bone than the IM group. Those hens also presented greater oviduct weight, greater hematocrit and greater serum concentration of total leukocytes, erythrocytes, eosinophils, monocytes and the hormones T4 and FSH, than the hens fed IM. The supplementation of AACM in laying hens' diets since one-day-old improves the productive performance from the beginning of egg output to peak production, which is justified by better development of bones and oviduct, hormone production and immune system support.

Background Malaria is one of the most critical global infectious diseases. Severe systemic inflammatory diseases, such as cerebral malaria, lead to the development of cognitive and behavioral alterations, such as learning disabilities and loss of memory capacity, as well as increased anxiety and depression. The consequences are profound and usually contribute to reduce the patient’s quality of life. There are no therapies to treat the neurological sequelae of cerebral malaria. Mesenchymal stromal cells (MSCs) may be an alternative, since they have been used as therapy for neurodegenerative diseases and traumatic lesions of the central nervous system. So far, no study has investigated the effects of MSC therapy on the blood-brain barrier, leukocyte rolling and adherence in the brain, and depression like-behavior in experimental cerebral malaria. Methods Male C57BL/6 mice were infected with Plasmodium berghei ANKA (PbA, 1 × 10 6 PbA-parasitized red blood cells, intraperitoneally). At day 6, PbA-infected animals received chloroquine (25 mg/kg orally for seven consecutive days) as the antimalarial treatment and were then randomized to receive MSCs (1 × 10 5 cells in 0.05 ml of saline/mouse) or saline (0.05 ml) intravenously. Parasitemia, clinical score, and survival rate were analyzed throughout the experiments. Evans blue assay was performed at 6, 7, and 15 days post-infection (dpi). Behavioral tests were performed at 5 and 15 dpi. Intravital microscopy experiments and brain-derived neurotrophic factor (BDNF) protein expression analyses were performed at 7 dpi, whereas inflammatory mediators were measured at 15 dpi. In vitro, endothelial cells were used to evaluate the effects of conditioned media derived from MSCs (CMMSC) on cell viability by lactate dehydrogenase (LDH) release. Results PbA-infected mice presented increased parasitemia, adherent leukocytes, blood-brain barrier permeability, and reduced BDNF protein levels, as well as depression-like behavior. MSCs mitigated behavioral alterations, restored BDNF and transforming growth factor (TGF)-β protein levels, and reduced blood-brain barrier dysfunction and leukocyte adhesion in the brain microvasculature. In a cultured endothelial cell line stimulated with heme, CMMSC reduced LDH release, suggesting a paracrine mechanism of action. Conclusion A single dose of MSCs as adjuvant therapy protected against vascular damage and improved depression-like behavior in mice that survived experimental cerebral malaria.

This study evaluated if the milk ring test (MRT) used to the brucellosis diagnostic at dairy herd level can be done in milk samples with conservative (Bronopol®) collected to the somatic cell count (SCC) that is used to mastitis monitoring. It were analyzed 38 bulk tank milk samples (BTMS) from 19 dairy herds artisanal cheese producers and 36 BTMS from a brucellosis free dairy herd, used as negative control (NC) and positive control (PC), when was added serum from vaccinated animals. The milk samples were collected in two different bottles (with or without Bronopol®). All the 38 BTMS from artisanal cheese producers (samples with and without Bronopol®) were non-reagent to MRT, as all NC. All PC were reagent to MRT, without interference of Bronopol®. Results showed that the milk sample used to SCC presented the proportion of agreement of results in all milk samples (100%), that is, false positive and false negative results were not observed. Results indicated that Bronopol® did not interfere with the color in the time of MRT reading suggesting that the same milk samples could be used to monitoring mastitis and brucellosis. RESUMO: O objetivo deste estudo foi avaliar se o teste do anel do leite (TAL), utilizado para o diagnóstico da brucelose em nível de rebanho leiteiro, pode ser realizado em amostras de leite com conservador (Bronopol®) coletado para contagem de células somáticas (CCS) que é utilizado para monitoramento de mastite. Foram analisadas 38 amostras de leite de tanques de expansão (ALTE) de 19 rebanhos leiteiros produtores de queijo artesanal e 36 ALTE de rebanho leiteiro livre de brucelose, utilizadas como controle negativo (CN) e controle positivo (CP), quando foi adicionado soro de animais vacinados. As amostras de leite foram coletadas em dois frascos diferentes (com e sem Bronopol®). Todos as 38 ALTE de produtores artesanais de queijo (amostras com e sem Bronopol®) foram não reagentes ao TAL, como todos os CN. Todos os CP foram reagentes ao TAL, sem interferência do Bronopol®. Os resultados mostraram que a amostra de leite utilizada para CCS apresentaram a proporção de concordância dos resultados em todas as amostras de leite (100%), ou seja, não foram observados resultados falso-positivos e falso-negativos. Os resultados indicam que o Bronopol® não interferiu na cor no momento da leitura do TAL, sugerindo que as mesmas amostras de leite poderiam ser usadas para monitorar mastite e brucelose.

The search for molecular markers to improve diagnosis, individualize treatment and predict behavior of tumors has been the focus of several studies. This study aimed to analyze homeobox gene expression profile in oral squamous cell carcinoma (OSCC) as well as to investigate whether some of these genes are relevant molecular markers of prognosis and/or tumor aggressiveness. Homeobox gene expression levels were assessed by microarrays and qRT-PCR in OSCC tissues and adjacent non-cancerous matched tissues (margin), as well as in OSCC cell lines. Analysis of microarray data revealed the expression of 147 homeobox genes, including one set of six at least 2-fold up-regulated, and another set of 34 at least 2-fold down-regulated homeobox genes in OSCC. After qRT-PCR assays, the three most up-regulated homeobox genes (HOXA5, HOXD10 and HOXD11) revealed higher and statistically significant expression levels in OSCC samples when compared to margins. Patients presenting lower expression of HOXA5 had poorer prognosis compared to those with higher expression (P=0.03). Additionally, the status of HOXA5, HOXD10 and HOXD11 expression levels in OSCC cell lines also showed a significant up-regulation when compared to normal oral keratinocytes. Results confirm the presence of three significantly upregulated (>4-fold) homeobox genes (HOXA5, HOXD10 and HOXD11) in OSCC that may play a significant role in the pathogenesis of these tumors. Moreover, since lower levels of HOXA5 predict poor prognosis, this gene may be a novel candidate for development of therapeutic strategies in OSCC.

Congenital generalized lipodystrophy (CGL) is a rare autosomal recessive disorder characterized by the absence of functional adipocytes resulting in ectopic lipid storage, metabolic disorders and early cardiovascular disease. Two-dimensional speckle-tracking (2D-STE) allows the detection of early abnormalities in myocardial function. We aimed to evaluate myocardial deformation in a large sample of CGL patients using 2D-STE. A cross-sectional study of 22 patients with CGL and 22 healthy subjects, matched for sex and age, was conducted from 2013 to 2018. All participants had undergone standard conventional echocardiography (ECHO) and 2D-STE. Determination of blood glucose, lipids, insulin, and leptin were performed in all CGL patients. In the CGL group the mean age was 14.6±10.7 years where 68.2% (n=15) were younger than 18 years old. All the patients had hypoleptinemia, 95.4% (21/22) low HDL-c, 86.4% (19/22) hypertriglyceridemia, 68.2% (15/22) diabetes, 50% (11/22) hepatic steatosis, 41% (9/22) insulin resistance, 41% (9/22) hypercholesterolemia, and 18.2% (4/22) hypertension. ECHO showed that 36.6% (8/22) of CGL patients presented diastolic dysfunction, 31.8% (7/22) left ventricular hypertrophy (LVH), 27.3% (6/22) increased left atrial volume index (LAVI), and 18.2% (4/22) increased left ventricular systolic diameter (LVDS) but normal ejection fraction (EF), whether using 2D-STE, 68.2% (15/22) of CGL patients showed abnormal global longitudinal strain (GLS) (p<0.01), and in almost LV segments. Positive association between abnormal GLS and A1c (r=0.57, p=0.005), glucose (r=0.5, p=0.018) and basal insulin (r= 0.69, p= 0.024), and negative association with leptin (r = -0.51, p = 0.005) were found in these patients. The 2D-STE revealed precocious left ventricular systolic dysfunction in a young CGL population with normal systolic function by ECHO. Early exposure to common metabolic abnormalities as insulin resistance, hyperglycemia, and hypoleptinemia must be involved in myocardial damage in these patients.