Lessons from a Multilaboratorial Task Force for Diagnosis of a Fatal Toxoplasmosis Outbreak in Captive Primates in Brazil.

Toxoplasmosis is an important zoonotic disease caused by the parasite Toxoplasma gondii and is especially fatal for neotropical primates. In Brazil, the Ministry of Health is responsible for national epizootic surveillance, but some diseases are still neglected. Here, we present an integrated investigation of an outbreak that occurred during the first year of the COVID-19 pandemic among eleven neotropical primates housed at a primatology center in Brazil. After presenting non-specific clinical signs, all animals died within four days. A wide range of pathogens were evaluated, and we successfully identified T. gondii as the causative agent within four days after necropsies. The liver was the most affected organ, presenting hemorrhage and hepatocellular necrosis. Tachyzoites and bradyzoite cysts were observed in histological examinations and immunohistochemistry in different organs; in addition, parasitic DNA was detected through PCR in blood samples from all specimens evaluated. A high prevalence of Escherichia coli was also observed, indicating sepsis. This case highlights some of the obstacles faced by the current Brazilian surveillance system. A diagnosis was obtained through the integrated action of researchers since investigation for toxoplasmosis is currently absent in national guidelines. An interdisciplinary investigation could be a possible model for future epizootic investigations in animals.

Trends of Infective Endocarditis at Two Teaching Hospitals: A 12-Year Retrospective Cohort Study in Rio de Janeiro, Brazil.

BACKGROUND: Despite advances in diagnosis and treatment, the incidence and mortality of infective endocarditis (IE) have increased in recent decades. Studies on the risk factors for mortality in endocarditis in Latin America are scarce. METHODS: This retrospective cohort study included 240 patients diagnosed with IE according to the modified Duke criteria who were admitted to two university hospitals in Rio de Janeiro, Brazil from January 2009 to June 2021. Poisson regression analysis was performed for trend tests. The multivariate Cox proportional hazards model was used to estimate the hazard ratio (HR) of predictors of in-hospital mortality. FINDINGS: The median age was 55 years (IQR: 39-66 years), 57% were male, and 41% had a Charlson comorbidity index (CCI) score > 3. Healthcare-associated infective endocarditis (54%), left-sided native valve IE (77.5%), and staphylococcal IE (26%) predominated. Overall, in-hospital mortality was 45.8%, and mortality was significantly higher in the following patients: aged ≥ 60 years (53%), CCI score ≥ 3 (60%), healthcare-associated infective endocarditis (HAIE) (53%), left-sided IE (51%), and enterococcal IE (67%). Poisson regression analysis showed no trend in in-hospital mortality per year. The adjusted multivariate model determined that age ≥ 60 years was an independent risk factor for in-hospital mortality (HR = 1.9; 95% CI 1.2-3.1; p = 0.008). INTERPRETATION: In this 12-year retrospective cohort, there was no evidence of an improvement in survival in patients with IE. Since older age is a risk factor for mortality, consensus is needed for the management of IE in this group of patients.

Seroprevalence of Hantavirus among Manual Cane Cutters and Epidemiological Aspects of HPS in Central Brazil.

Hantavirus pulmonary syndrome (HPS) is a rodent-borne zoonotic disease that is endemic throughout the Americas. Agricultural activities increase exposure to wild rodents, especially for sugarcane cutters. We carried out a survey of the epidemiological aspects of HPS and investigated the prevalence of hantavirus infection in the sugarcane cutter population from different localities in the Brazilian Midwest region. We conducted a retrospective study of all confirmed HPS cases in the state of Goiás reported to the National HPS surveillance system between 2007 and 2017, along with a seroepidemiological study in a population of sugarcane cutters working in Goiás state in 2016, using the anti-hantavirus (Andes) ELISA IgG. A total of 634 serum samples from cane cutters were tested for hantavirus antibodies, with 44 (6.9%) being IgG-reactive according to ELISA. The destination of garbage was the only statistically significant variable (p = 0.03) related to the detection of hantavirus IgG (p < 0.05). We described the epidemiological profile of reported hantavirus cases in Goiás-a highly endemic area for HPS, and where the seroepidemiological study was conducted. Our results increase our knowledge about hantavirus infections in Brazil and highlight the vulnerability of sugarcane cutters to a highly lethal disease that, to date, has no specific treatment or vaccination.

Emergence of melioidosis in Brazil: a case series.

BACKGROUND: Melioidosis is a serious disease caused by the bacterium Burkholderia pseudomallei which affects humans and animals. It results in a wide spectrum of clinical manifestations, mainly in the respiratory tract, progressing to septic shock and death. CASE PRESENTATION: Herein, we present a series of seven patients (median age: 41 years) with confirmed melioidosis, treated at a public hospital in Piauí State, Brazil between 2019 and 2021. The most common clinical presentations were fever, cough, pneumonia, and abdominal pain. The mean duration of antibacterial therapy with 1 g of meropenem was 28.6 ± 1.1 days. Six patients recovered and one died. The mean hospitalization time was 51.0 ± 39.2 days. CONCLUSIONS: Melioidosis is an emerging infectious disease in Brazil. Health professionals in endemic areas need to be aware of the clinical presentation and appropriate clinical management of the disease because early diagnosis and early initiation of antibiotic therapy can be life-saving.

Case Report: Hantavirus Cardiopulmonary Syndrome Diagnostic in the Face of the COVID-19 Pandemic.

Hantavirus cardiopulmonary syndrome (HCPS) is an emerging rodent-borne disease in the Americas. The most common initial symptoms of HCPS are similar to those of COVID-19 and other respiratory infections that evolve rapidly to respiratory failure, resulting from pulmonary edema and shock in about 40% of cases. We describe a fatal case of HCPS in a 24-year-old man who was hospitalized with fever, hemoconcentration, thrombocytopenia, leukocytosis, dry cough and a bilateral diffuse alveolar pulmonary infiltrate during the emergence of the COVID-19 pandemic in Brazil. HCPS needs to be ruled out in patients with clinical manifestations compatible with respiratory infections such as influenza and COVID-19.

Seroprevalence estimate and risk factors for Coxiella burnetii infections among humans in a highly urbanised Brazilian state.

BACKGROUND: Q fever is among the top 13 global priority zoonoses, however, it is still neglected and under-reported in most of the world, including Brazil. Thus, we evaluated the seroprevalence of and the risk factors for Coxiella burnetii infections in humans from Minas Gerais, a highly urbanised Brazilian state. METHODS: Coxiella burnetii was searched for patient samples (n=437), which were suspected of then later confirmed as negative for dengue fever, by the indirect immunofluorescence technique and real-time PCR. Risk factors for infections and spatial clusters for both C. burnetii-seropositive individuals and livestock concentration were evaluated. RESULTS: We found that 21 samples (4.8%; 95% CI 3.0 to 7.2%) were reactive for at least one class of anti-C. burnetii antibodies (titer of ≥64), with rural residence (p=0.036) being a risk factor. Also, two spatial clusters of seropositivity were found within a significant area by Scan, and a probable relationship between the Scan result and the livestock concentration by area was found. CONCLUSIONS: Seropositive individuals were associated with rural residence, with a likely relationship with the livestock concentration. Thus, this study establishes baseline figures for C. burnetii seroprevalence in humans in a state of Brazil, allowing the monitoring of trends and setting of control targets, as well as more representative longitudinal and risk analysis studies.

Identification of Immunogenic Linear B-Cell Epitopes in C. burnetii Outer Membrane Proteins Using Immunoinformatics Approaches Reveals Potential Targets of Persistent Infections.

Coxiella burnetii is a global, highly infectious intracellular bacterium, able to infect a wide range of hosts and to persist for months in the environment. It is the etiological agent of Q fever-a zoonosis of global priority. Currently, there are no national surveillance data on C. burnetii's seroprevalence for any South American country, reinforcing the necessity of developing novel and inexpensive serological tools to monitor the prevalence of infections among humans and animals-especially cattle, goats, and sheep. In this study, we used immunoinformatics and computational biology tools to predict specific linear B-cell epitopes in three C. burnetii outer membrane proteins: OMP-H (CBU_0612), Com-1 (CBU_1910), and OMP-P1 (CBU_0311). Furthermore, predicted epitopes were tested by ELISA, as synthetic peptides, against samples of patients reactive to C. burnetii in indirect immunofluorescence assay, in order to evaluate their natural immunogenicity. In this way, two linear B-cell epitopes were identified in each studied protein (OMP-H(51-59), OMP-H(91-106), Com-1(57-76), Com-1(191-206), OMP-P1(197-209), and OMP-P1(215-227)); all of them were confirmed as naturally immunogenic by the presence of specific antibodies in 77% of studied patients against at least one of the identified epitopes. Remarkably, a higher frequency of endocarditis cases was observed among patients who presented an intense humoral response to OMP-H and Com-1 epitopes. These data confirm that immunoinformatics applied to the identification of specific B-cell epitopes can be an effective strategy to improve and accelerate the development of surveillance tools against neglected diseases.

Orthohantavirus Survey in Indigenous Lands in a Savannah-Like Biome, Brazil.

In Brazil, the first confirmed cases of hantavirus cardiopulmonary syndrome in Indigenous populations occurred in 2001. The purpose of this study was to determine the seroprevalence of orthohantavirus infections in the Utiariti Indigenous land located in the southeastern region of the Brazilian Amazon. In December 2014 and 2015, a survey was conducted using an enzyme-linked immunosorbent assay in nine villages belonging to the Haliti-Paresí Indigenous communities. A total of 301 participants were enrolled in the study. Of the two study cohorts, the one from 2014 showed a prevalence of 12.4%, whereas the one from 2015 had a serum prevalence of 13.4%. Analysis of the paired samples of 110 Indigenous people who participated in both stages of the study enabled identification of four individuals who had seroconverted during the study period. Identifying the circulation of orthohantaviruses in the Utiariti Indigenous land highlights a serious public health problem in viral expansion and highlights the need to implement preventive measures appropriate to the sociocultural reality of these communities.

Human Q Fever on the Guiana Shield and Brazil: Recent Findings and Remaining Questions.

PURPOSE OF REVIEW: In this review, we report on the state of knowledge about human Q fever in Brazil and on the Guiana Shield, an Amazonian region located in northeastern South America. There is a contrast between French Guiana, where the incidence of this disease is the highest in the world, and other countries where this disease is practically non-existent. RECENT FINDINGS: Recent findings are essentially in French Guiana where a unique strain MST17 has been identified; it is probably more virulent than those usually found with a particularly marked pulmonary tropism, a mysterious animal reservoir, a geographical distribution that raises questions. SUMMARY: Q fever is a bacterial zoonosis due to Coxiella burnetii that has been reported worldwide. On the Guiana Shield, a region mostly covered by Amazonian forest, which encompasses the Venezuelan State of Bolivar, Guyana, Suriname, French Guiana, and the Brazilian State of Amapá, the situation is very heterogeneous. While French Guiana is the region reporting the highest incidence of this disease in the world, with a single infecting clone (MST 117) and a unique epidemiological cycle, it has hardly ever been reported in other countries in the region. This absence of cases raises many questions and is probably due to massive under-diagnosis. Studies should estimate comprehensively the true burden of this disease in the region.

Frequency of co-seropositivities for certain pathogens and their relationship with clinical and histopathological changes and parasite load in dogs infected with Leishmania infantum.

In canine leishmaniosis caused by the protozoan Leishmania infantum, little is known about how co-infections with or co-seropositivities for other pathogens can influence aggravation of this disease. Therefore, the objectives of this study were to evaluate the frequency of co-infections with or co-seropositivities for certain pathogens in dogs seropositive for L. infantum and their relationship with clinical signs, histological changes and L. infantum load. Sixty-six L. infantum-seropositive dogs were submitted to clinical examination, collection of blood and bone marrow, culling, and necropsy. Antibodies against Anaplasma spp., Borrelia burgdorferi sensu lato, Ehrlichia spp. and Toxoplasma gondii and Dirofilaria immitis antigens were investigated in serum. Samples from different tissues were submitted to histopathology and immunohistochemistry for the detection of Leishmania spp. and T. gondii. Quantitative real-time PCR was used to assess the L. infantum load in spleen samples. For detection of Coxiella burnetii, conventional PCR and nested PCR were performed using bone marrow samples. All 66 dogs tested positive for L. infantum by qPCR and/or culture. Fifty dogs (76%) were co-seropositive for at least one pathogen: T. gondii (59%), Ehrlichia spp., (41%), and Anaplasma spp. (18%). Clinical signs were observed in 15 (94%) dogs monoinfected with L. infantum and in 45 (90%) dogs co-seropositive for certain pathogens. The L. infantum load in spleen and skin did not differ significantly between monoinfected and co-seropositive dogs. The number of inflammatory cells was higher in the spleen, lung and mammary gland of co-seropositive dogs and in the mitral valve of monoinfected dogs. These results suggest that dogs infected with L. infantum and co-seropositive for certain pathogens are common in the region studied. However, co-seropositivities for certain pathogens did not aggravate clinical signs or L. infantum load, although they were associated with a more intense inflammatory reaction in some organs.

Molecular and Serological Survey of the Cat-Scratch Disease Agent (Bartonella henselae) in Free-Ranging Leopardus geoffroyi and Leopardus wiedii (Carnivora: Felidae) From Pampa Biome, Brazil.

The genus Bartonella comprises emerging bacteria that affect humans and other mammals worldwide. Felids represent an important reservoir for several Bartonella species. Domestic cats are the main reservoir of Bartonella henselae, the agent of cat scratch disease (CSD). It can be transmitted directly by scratches and bites from infected cats and via cat fleas. This study aims to investigate the circulation of Bartonella spp. in free-ranging Neotropical wild felids from Southern Brazil using serological and molecular methods. In this study, 53 live-trapped free-ranging wild felids were sampled, 39 Leopardus geoffroyi and 14 Leopardus wiedii, from five municipalities in the Rio Grande, do Sul state, southern Brazil. All captured animals were clinically healthy. Two blood samples of L. geoffroyi were positive, by PCR, for the presence of B. henselae DNA. Conversely, none of L. wiedii blood samples were positive when tested using PCR. Indirect immunofluorescence assay (IFA) showed that 28% of serum samples of wild felids were reactive (seropositive) for B. henselae by immunofluorescence, with titers ranging from 64 to 256. The results presented here provide the first evidence of a Bartonella-enzootic cycle involving L. geoffroyi and L. wiedii, which may account for the spillover of the emerging zoonotic pathogen B. henselae for the indigenous fauna in Southern Brazil.

MicroRNAs and Mammarenaviruses: Modulating Cellular Metabolism.

Mammarenaviruses are a diverse genus of emerging viruses that include several causative agents of severe viral hemorrhagic fevers with high mortality in humans. Although these viruses share many similarities, important differences with regard to pathogenicity, type of immune response, and molecular mechanisms during virus infection are different between and within New World and Old World viral infections. Viruses rely exclusively on the host cellular machinery to translate their genome, and therefore to replicate and propagate. miRNAs are the crucial factor in diverse biological processes such as antiviral defense, oncogenesis, and cell development. The viral infection can exert a profound impact on the cellular miRNA expression profile, and numerous RNA viruses have been reported to interact directly with cellular miRNAs and/or to use these miRNAs to augment their replication potential. Our present study indicates that mammarenavirus infection induces metabolic reprogramming of host cells, probably manipulating cellular microRNAs. A number of metabolic pathways, including valine, leucine, and isoleucine biosynthesis, d-Glutamine and d-glutamate metabolism, thiamine metabolism, and pools of several amino acids were impacted by the predicted miRNAs that would no longer regulate these pathways. A deeper understanding of mechanisms by which mammarenaviruses handle these signaling pathways is critical for understanding the virus/host interactions and potential diagnostic and therapeutic targets, through the inhibition of specific pathologic metabolic pathways.

Evaluation of rickettsial infection in free-range capybaras (Hydrochoerus hydrochaeris Linnaeus, 1766) (Rodentia: Caviidae) and ticks (Acari: Ixodidae) in the Western Amazon, Brazil.

Capybaras (Hydrochoerus hydrochaeris Linnaeus, 1766) (Rodentia: Caviidae) are important hosts of Amblyomma ticks (Acari: Ixodidae), which in turn can transmit rickettsiae to humans and animals. However, there is a scarcity of studies about the tick fauna and rickettsial infection in the Amazon region. The present study evaluated rickettsial infection in capybaras and ticks in different areas of the municipality of Rio Branco, state of Acre, in the Western Brazilian Amazon, where rickettsiosis has never been reported. Blood sera from 43 capybaras from four localities in Rio Branco were tested by indirect immunofluorescence assay using Rickettsia rickettsii antigens. Ticks were collected from capybaras and from vegetation as well. Ticks were taxonomically identified to the species level and some of them were tested by PCR, targeting a fragment of the rickettsial gltA gene. Additionally, ticks were tested for bacteria from the genus Borrelia and family Anaplasmatacae. All capybaras submitted to the serological examination were considered non-reactive to R. rickettsii. A total of 410 ticks were collected directly from the capybaras. Amblyomma dubitatum Neumann, 1899 was the most abundant species (82.4%), followed by Amblyomma naponense (Packard, 1869) (14.3%), Amblyomma humerale Koch, 1844 (0.7%), Amblyomma pacae Aragão, 1911 (0.4%), Amblyomma rotundatum Koch 1844 (0.2%) and Amblyomma sp. (1.7%). From the environment 262 ticks were collected: Rhipicephalus microplus (Canestrini, 1888) (88.9%), Dermacentor nitens Neumann, 1897 (9.9%), Amblyomma varium Koch, 1844 (0.7%) and A. rotundatum (0.3%). With the exception of A. humerale, A. rotundatum and R. microplus, all other species are reported here for the first time in the state. Some of the ticks sampled (N = 317) were tested by molecular methods for infection by Rickettsia spp. Rickettsia bellii was identified infecting A. dubitatum and A. rotundatum, while Rickettsia amblyommatis only was found infecting A. humerale and Rickettsia sp. strain Tapirapé was found in A. naponense. This is the first detection of R. bellii and Rickettsia sp. strain Tapirapé in Acre. No Borrelia or Anaplasmataceae were found in the tested ticks. These results add relevant knowledge about the Rickettsia spp. and the acarological fauna in the region of the Western Amazon, and are essential for the maintenance of vigilance about possible pathogens that occur in the state and determination of the risks that they pose to humans and animals that inhabit the region.

Aporé virus, a novel mammarenavirus (Bunyavirales: Arenaviridae) related to highly pathogenic virus from South America.

Here, we report the complete genome sequence of the Aporé virus (Bunyavirales: Arenaviridae), obtained from a wild rodent Oligoryzomys mattogrossae captured in Mato Grosso do Sul state, Brazil. The genome of this virus showed strong similarity to highly pathogenic mammarenavirus from South America.

First description of Theiler's disease-associated virus infection and epidemiological investigation of equine pegivirus and equine hepacivirus coinfection in Brazil.

Recent advances in the study of equine pegivirus (EPgV), Theiler's disease-associated virus (TDAV) and equine hepacivirus (EqHV) highlight their importance to veterinary and human health. To gain some insight into virus distribution, possible risk factors, presence of liver damage and genetic variability of these viruses in Brazil, we performed a cross-sectional study of EPgV and TDAV infections using a simultaneous detection assay, and assessed EqHV coinfection in different horse cohorts. Of the 500 serum samples screened, TDAV, EPgV and EPgV-EqHV were present in 1.6%, 14.2% and 18.3%, respectively. EPgV-positive horses were present in four Brazilian states: Espírito Santo, Mato Grosso do Sul, Minas Gerais and Rio de Janeiro. Serum biochemical alterations were present in 40.4% of EPgV-infected horses, two of them presenting current liver injury. Chance of infection was 2.7 times higher in horses ≤5 years old (p = 0.0008) and 4.9 times higher in horses raised under intensive production systems (p = 0.0009). EPgV-EqHV coinfection was 75% less likely in horses older than 5 years comparatively to those with ≤5 years old (p = 0.047). TDAV-positive animals were detected in different horse categories without biochemical alteration. Nucleotide sequences were highly conserved among isolates from this study and previous field and commercial product isolates (≥88% identity). Tree topology revealed the formation of two clades (pp = 1) for both EPgV and TDAV NS3 partial sequences. In conclusion, the widespread presence of EPgV-RNA suggests an enzootic infection with subclinical viremia in Brazil. Horse management can influence virus spread. This first report of TDAV-infected horses outside the USA reveals the existence of subclinical viremic horses in distant geographical regions. EPgV and TDAV have similar circulating isolates worldwide. These findings contribute to global efforts to understand the epidemiology and pathogenesis of these equine viruses.

Detected microorganisms and new geographic records of Ornithodoros rietcorreai (Acari: Argasidae) from northern Brazil.

Reliable data on distributional ranges of soft ticks (Argasidae) and assessments of putative tick-borne agents enhance the understanding on tick-associated microorganisms. A total of 96 ticks morphologicaly and molecularly identified as Ornithodoros rietcorreai were collected in Tocantins State, Brazil, using Noireau traps with living bait as CO2 source. Ninety-six ticks (54 nymphs, 32 males, 10 females) with different engorgement degrees were collected. Fourty-seven (48.9%) of them were individually screened by PCR for detecting bacteria of Anaplasmataceae family and genera Rickettsia, and Borrelia. The presence of protozoans of the genus Babesia was assessed as well. Fourty seven ticks were submitted to analysis. Nine ticks (19.1%) yielded sequences for gltA and htrA genes most identical with a series of endosymbiont rickettsiae and Rickettsia bellii, respectively. Upon two ticks (4.2%) we retrieved DNA of a potential new Wolbachia sp., and DNA of a putative novel Hepatozoon was characterized from three (6.4%) specimens. No DNA of Babesia or Borrelia was detected. Remarkably, amplicons of unidentified eukaryotic organisms, most closely related with apicomplexans but also with dinoflagellates (91% of identity after BLAST analyses), were recovered from two ticks (4.2%) using primers designed for Babesia 18S rRNA gene. Our records expand the distribution of O. rietcorreai into Brazilian Cerrado biome and introduce the occurrence of microorganisms in this tick species.

The mystery of the phylogeographic structural pattern in rodent-borne hantaviruses.

Hantaviruses (order Bunyavirales, family Hantaviridae) are important zoonotic pathogens. Because of the great diversity of their reservoir hosts, hantaviruses are excellent models to evaluate the dynamics of virus-host co-evolution. To understand the mechanisms behind the evolutionary history of hantaviruses through virus-reservoir interactions, it is important to know how the radiation and diversity of hantaviruses occurred. In this paper, we evaluate the pattern of hantavirus diversification based on a complete S segment representing major groups of hantaviruses found in the Americas. Phylogenetic analyses revealed a high degree of phylogeographic structure and a surprising pattern of geographical distribution of New World hantaviruses. The available data suggest that hantaviruses related to the Arvicolinae rodent subfamily in North America probably emerged and initially adapted from a shared common ancestor of the Tula virus. The first clade of hantaviruses associated with Neotominae occupied a stem lineage, especially those that emerged in Central America or Mexico. Hantaviruses from Central America and Mexico found in Neotominae rodents spread northward and probably gave rise to the first phylogroup of hantaviruses associated with Sigmodontinae in North America. Two preferential host-switching transmissions in hantaviruses apparently gave rise to two different paraphyletic group in Neotominae and Sigmodontinae. Our study supports a probable epicenter of diversification in Central America and/or Mexico for hantaviruses related to both the Neotominae and Sigmodontinae subfamilies.

What is the minimum length of gltA gene required for phylogenetic analyzes in Bartonella?

Bartonellosis is an emerging serious diseases of zoonotic relevance caused by an expanding number of recently discovered species of Bartonella. Many studies use partial gltA gene sequencing as the method of choice for species attribution, and a plethora of studies have utilized only this gene to describe Bartonella diversity. We observe a lack of correspondence between the phylogenies constructed using the complete gltA gene sequences and the small gene fragment usually used in phylogenetic analyzes. Everything indicates that important changes occurred by using larger fragments. Therefore, it is of great importance to define a minimum gltA fragment length to be used in future phylogenetic analyzes, besides the use of other genes.

Aporé virus, a novel mammarenavirus (Bunyavirales: Arenaviridae) related to highly pathogenic virus from South America

Here, we report the complete genome sequence of the Aporé virus (Bunyavirales: Arenaviridae), obtained from a wild rodent Oligoryzomys mattogrossae captured in Mato Grosso do Sul state, Brazil. The genome of this virus showed strong similarity to highly pathogenic mammarenavirus from South America.