Molecular detection of Toxoplasma gondii and Neospora caninum in neotropical primates.

BACKGROUND: Toxoplasma gondii and Neospora caninum infections in primates are potentially fatal and directly impact the conservation of these animals and public health. MATERIALS AND METHODS: A total of 38 blood/clot samples collected from free-living and captive neotropical primates undergoing clinical care or found dead by environmental authorities in the Mato Grosso State, Brazil, were analyzed by PCR for DNA detection of T. gondii and N. caninum. Furthermore, eight animals were submitted to immunohistochemistry for the detection of T. gondii. RESULTS: DNA of T. gondii and N. caninum was amplified in 11 (28.95%) 10 (26.32%) of samples analyzed, respectively. Coinfection was observed in three individuals. One animal returned a positive result in the immunohistochemistry for the detection of T. gondii. CONCLUSION: These findings reflect a concern for the conservation of these animals, as the pathogen-host interaction is unpredictable and infections by these protozoa can lead to animal mortality, which has a substantial impact on endangered species.

Domestic Birds as Source of Cryptococcus deuterogattii (AFLP6/VGII): Potential Risk for Cryptococcosis.

Cryptococcosis is an infection caused by encapsulated basidiomycetous yeasts belonging to the Cryptococcus neoformans/Cryptococcus gattii species complexes. It is acquired through inhalation of infectious propagules, often resulting in meningitis and meningoencephalitis. The ecological niche of these agents is a wide variety of trees species, as well as pigeon, parrot and passerine excreta. The objective of this study was to isolate Cryptococcus yeasts from excreta of commercially traded parrots and passerines. The 237 samples were collected between October 2018 and April 2019 and processed using conventional methodologies. Nineteen colonies with a dark brown phenotype, caused by phenol oxidase activity, were isolated, suggesting the presence of pathogenic Cryptococcus yeasts. All isolates tested positive for urease activity. URA5-RFLP fingerprinting identified 14 isolates (68.4%) as C. neoformans (genotype AFLP1/VNI) and 5 (26.3%) as C. deuterogattii (genotype AFLP6/VGII). Multi-locus sequence typing was applied to investigate the relatedness of the C. deuterogattii isolates with those collected globally, showing that those originating from bird-excreta were genetically indistinguishable from some clinical isolates collected during the past two decades.

Pasteurella canis infection in a non-human primate black-tailed marmoset (Mico melanurus)-A case report.

This study reports the pathological and microbiological findings of pneumonia in a Mico melanurus caused by Pasteurella canis, confirmed for molecular analyses. It demonstrated the importance that wild species represent in the epidemiology of pasteurellosis in anthropic environments, when inserted into urban areas.

Ruminal Microbial Populations and Fermentation Characteristics in Beef Cattle Grazing Tropical Forage in Dry Season and Supplemented with Different Protein Levels.

We tested the hypothesis that supplementation with protein improves fermentation parameters without damaging the rumen microbial populations of beef cattle grazing Urochloa brizantha cv. Marandu during the dry season. Four rumen-cannulated Nellore bulls (571 ± 31 kg of body weight) were used in a 4 × 4 Latin square design. The treatments were not supplemented with concentrate (only free-choice mineral salt ad libitum) and supplemented (supplements with low-LPSU, medium-MPS, and high protein supplement-HPS), supplying 155, 515, and 875 g/animal/day of crude protein (CP), respectively. The abundance of each target taxon was calculated as a fraction of the total 16S rRNA gene copies in the samples, using taxon-specific and domain bacteria primers. There was no difference (P > 0.05) across treatments for intakes of dry matter (DM), forage and neutral detergent fiber (NDF), digestibility of DM and NDF, and ruminal pH. Animals supplemented with concentrate had greater (P < 0.05) intakes and digestibility of CP, ether extract and non-fibrous carbohydrate contents of the substrates (EE + NFC), and ruminal ammonia nitrogen (RAN) compared to control. Bulls that received only mineral salt had lower proportions of Butyrivibrio fibrisolvens and had greater (P < 0.05) proportions of Fibrobacter succinogenes, Ruminococcus albus, Ruminococcus flavefaciens, Methanogen archaea than bulls supplemented with concentrate. The MPS animals had greater (P < 0.05) intake and digestibility of CP, RAN concentration, and had lower (P < 0.05) proportions of Fibrobacter succinogenes, Ruminococcus flavefaciens, and Selenomonas ruminantium than LPSU animals. The HPS provided higher (P < 0.05) intake of CP, RAN and proportion of Ruminococcus albus when compared with MPS. In conclusion, supply of 515 g/animal/day of protein via supplement provides better ruminal conditions for the growth of cellulolytic bacteria of bulls on pasture during dry season.

Genetic Diversity of the Cryptococcus gattii Species Complex in Mato Grosso State, Brazil.

Cryptococcosis is caused by fungi of the genus Cryptococcus. Owing to its importance, this study aimed to analyze the genetic diversity of C. gattii isolates from animals, humans, and the environment in Mato Grosso State (MT), Brazil, during November 2010-December 2017. All isolates of the C. gattii species complex were subjected to molecular genotyping via Restriction Fragment Length Polymorphism (PCR-RFLP) and Multi-locus Sequence Typing (MLST). PCR-RFLP analysis revealed that 21 isolates presented the genotype VGII, which is considered the most common and virulent genotype globally among. MLST analysis revealed the presence of 14 sequence types (STs), of which 5 are considered new genotypes. Clonal Complex (CC) CC182 (n = 5; 23,80%) and CC309 (n = 3; 14,28%) were the most frequent. CC distribution in relation to origin revealed that three CCs were found in animals with a predominance of CC182 (66,66%), while nine were found in humans, and two CCs were found in the environment. Extensive genetic variability was observed among the isolates in the State of Mato Grosso. STs belonging to the already described clonal complexes (CC) indicate the global expansion and adaptation of isolates in several other countries. Therefore, detection of clonal complexes and STs already described in other regions and the occurrence of new STs in the present study help further the current understanding of the geographic dispersion and genetic origin of the C. gattii species complex.

Identification of Pasteurella multocida transcribed genes in porcine lungs through RNAseq.

Pasteurella multocida is one of the most important pathogen that causes pneumonia in swine. Although several virulence factors are known, the pathogenesis of this bacterium is not well-studied. Therefore, to study the pathogenesis of P. multocida infection in porcine lung, next-generation RNA sequencing was used to compare the transcriptomes of P. multocida grown in vivo and in vitro, respectively. After P. multocida infection a total of 704 genes were expressed in vitro, 1422 genes were expressed in vivo, and 237 genes were differentially expressed based on statistical analyses, padj of ≤0.1. Genes encoding ribosomal proteins or other products that function in the regulation of transcription and translation were downregulated, whereas genes whose products affected cellular processes (protein transport and RNA degradation) and metabolic pathways, such as those of amino acid metabolism and nucleotide metabolism, were upregulated in vitro compared with in vivo. This study shows that differentially expressed genes in P. multocida regulate pathways that operate during stress, iron capture, heat shock, and nitrogen regulation. However, extensive investigation of the pathogenic mechanism of P. multocida is still required.

Detection of noroviruses in free-ranging jaguars (Panthera onca) in the Pantanal, Mato Grosso, Brazil.

Nine free-ranging jaguars (Panthera onca) were captured, and rectal swabs were collected in the Pantanal of Cáceres, Mato Grosso, Brazil. Reverse transcription polymerase chain reaction specific for noroviruses was performed. Six jaguars (66.6%) tested positive for norovirus genotype GII.11.

Quantitative qPCR Analysis of Ruminal Microorganisms in Beef Cattle Grazing in Pastures in the Rainy Season and Supplemented with Different Protein Levels.

We tested the hypothesis that supplementation with three protein levels improves fermentation parameters without changing the rumen microbial population of grazing beef cattle in the rainy season. Four rumen-cannulated Nellore bulls (432 ± 21 kg of body weight) were used in a 4 × 4 Latin square design with four supplements and four experimental periods of 21 days each. The treatments were mineral supplement (ad libitum) and supplements with low, medium (MPS), and high protein supplement (HPS), supplying 106, 408, and 601 g/day of CP, respectively. The abundance of each target taxon was calculated as a fraction of the total 16S rRNA gene copies in the samples, using taxon-specific and domain bacteria primers. Supplemented animals showed lower (P < 0.05) proportions of Ruminococcus flavefaciens and greater (P < 0.05) proportions of Ruminococcus albus and Butyrivibrio fibrisolvens than animals that received only the mineral supplement. The HPS supplement resulted in higher (P < 0.05) proportions of Fibrobacter succinogenes, R. flavefaciens, and B. fibrisolvens and lower (P < 0.05) proportions of R. albus than the MPS supplement. Based on our results, high protein supplementation improves the ruminal conditions and facilitates the growth of cellulolytic bacteria in the rumen of bulls on pastures during the rainy season.

Experimental infection in gerbils by Conidiobolus lamprauges.

Conidiobolomycosis is an emerging entomophthoramycosis caused by fungi Conidiobolus spp. Animal models are essential for the study of infectious disease in various areas such as pathogenesis, diagnostic methods, treatment and prevention. There is not currently an animal model for conidiobolomycosis. The aim of this study was to create an experimental infection protocol for Conidiobolus lamprauges in gerbils (Meriones unguiculatus). The study animals were randomly divided into four groups of four animals: immunosuppressed with cyclophosphamide (CPA) and infected with C. lamprauges (G1), immunocompetent and infected with C. lamprauges (G2), immunosuppressed with CPA (G3), and an immunocompetent control group (G4). Clinical signs were observed only in G1 animals, where the mortality rate reached 75% by day 7 after infection (AI) with a median survival of 2 days. C. lamprauges was detected only in G1, both by PCR and by isolation. Necropsies of the G1 animals showed lesions in the nasal cavity and lung tissue. These lesions were characterized by polymorphonuclear infiltrate cells and by the presence of hyphal structures under silver staining. This animal model will be useful for further investigation of diseases caused by C. lamprauges, particularly of those associated with immunosuppression factors in naturally occurring animal infections.

Presence of Ureaplasma diversum in the genital tracts of female dairy cattle in Mato Grosso State, Brazil.

Ureaplasma diversum infection in bovine females may result in various reproductive problems, including granular vulvovaginitis, abortion, weak calves, salpingitis, and spontaneous abortion. The presence of U. diversum in a dairy bovine population from midwestern Brazil has not been established. The aim of this study was to determine whether U. diversum was present in dairy cattle from midwestern Brazil using polymerase chain reaction (PCR). Vulvovaginal mucus was analyzed from 203 cows located in six municipalities in the north region of Mato Grosso State, Brazil. A total of 25% of dairy cows with vulvovaginitis were positive for U. diversum. The factors evaluated were included in a multivariable logistic regression model with the presence of at least one positive cow in the herd serving as the dependent variable. Three variables were significantly associated with a U. diversum-positive PCR and were included in the final multivariable model: number of parities, vulvar lesions, and reproductive problems. For each new parity, the chance of U. diversum infection decreased 0.03-fold, indicating that cows with the highest number of parities were more protected. The presence of vulvar lesions was increased 17.6-fold in females positive for U. diversum, suggesting that this bacterium could be related to the red granular lesions in the vulvar mucosa, whereas reproductive problems were increased 7.6-fold. However, further investigations should be conducted to ascertain the effects of U. diversum in association with other mycoplasma species in the herds studied.

Cryptococcus gattii VGII in a Plathymenia reticulata hollow in Cuiabá, Mato Grosso, Brazil.

Little is known about the ecology of agents of cryptococcosis in Mato Grosso, without any data regarding to the sources of both agents in the environment. This study aimed to investigate Cryptococcus gattii and Cryptococcus neoformans associated with decay in tree hollows within the urban area of three different cities of Mato Grosso. Seventy-two environmental samples collected from 72 living trees in the cities of Cuiabá, Várzea Grande and Chapada dos Guimarães were sampled and analysed. One tree (Plathymenia reticulata, Leguminosae) in the city of Cuiabá yielded 19 colonies identified as C. gattii molecular type VGII. The isolation of C. gattii VGII in the downtown city of Cuiabá is important because it fits in the Northern Macroregion, suggesting expanding and urbanisation of this genotype in different Brazilian cities.

Phlebotomine fauna, natural infection rate and feeding habits of Lutzomyia cruzi in Jaciara, state of Mato Grosso, Brazil.

Visceral leishmaniasis (VL) in Brazil is transmitted by the phlebotomine Lutzomyia longipalpis and in some midwestern regions by Lutzomyia cruzi. Studies of the phlebotomine fauna, feeding habits and natural infection rate by Leishmania contribute to increased understanding of the epidemiological chain of leishmaniases and their vectorial capacity. Collections were performed in Jaciara, state of Mato Grosso from 2010-2013, during which time 2,011 phlebotomines (23 species) were captured (68.70% Lu. cruzi and 20.52% Lutzomyia whitmani). Lu. cruzi females were identified by observing the shapes of the cibarium (a portion of the mouthpart) and spermatheca, from which samples were obtained for polymerase chain reaction to determine the rates of natural infection. Engorged phlebotomines were assessed to identify the blood-meal host by ELISA. A moderate correlation was discovered between the number of Lu. cruzi and the temperature and the minimum rate of infection was 6.10%. Twenty-two females were reactive to the antisera of bird (28%), dog (3.30%) and skunk (1.60%). We conclude that Lu. cruzi and Lu. whitmani have adapted to the urban environment in this region and that Lu. cruzi is the most likely vector of VL in Jaciara. Moreover, maintenance of Leishmania in the environment is likely aided by the presence of birds and domestic and synanthropic animals.

Ureaplasma diversum as a cause of pustular vulvovaginitis in bovine females in Vale Guapore, Mato Grosso State, Brazil.

Ureaplasma diversum has been associated with various reproductive problems in cattle that include granular vulvovaginitis, weak calves, and abortion. This study was conducted in a beef herd situated in the Middle-West region of Brazil, and the objectives were to verify the presence of U. diversum and to elucidate its possible relationships with independent variables in this bovine herd population. A total of 134 vaginal mucous swabs were taken for polymerase chain reaction (PCR). Of these, 51 (38 %) were PCR positive for U. diversum. Of the 58 heifers with vulvovaginal lesions characterized by hyperemia, granulated lesions, and edema distributed throughout the vulvar mucosa, 37 (64 %) were U. diversum positive; of the 76 heifers without reproductive lesions, 14 (18 %) were U. diversum positive. All tested samples were negative for bovine herpesvirus 1 (BoHV-1). Multivariate logistic regression revealed that the following two variables were significantly associated with the presence of U. diversum: the presence of vulvar lesions (p = 0.001) and the presence of a progesterone (P4) device (p = 0.001). These findings indicate that U. diversum should be considered a pathogen that is associated with pustular vulvovaginitis in heifers from the Mato Grosso state and that additional studies of the risk factors associated with intravaginal P4 device transmission should be performed.

Whole-genome sequencing analysis of multidrug-resistant Serratia marcescens isolates in an intensive care unit in Brazil.

INTRODUCTION: Serratia marcescens is an opportunistic pathogen found ubiquitously in the environment and associated with a wide range of nosocomial infections. This multidrug-resistant bacterium has been a cause of concern for hospitals and healthcare facilities due to its ability to spread rapidly and cause outbreaks. Next generation sequencing genotyping of bacterial isolates has proven to be a valuable tool for tracking the spread and transmission of nosocomial infections. This has allowed for the identification of outbreaks and transmission chains, as well as determining whether cases are due to endogenous or exogenous sources. Evidence of nosocomial transmission has been gathered through genotyping methods. The aim of this study was to investigate the genetic diversity of carbapenemase-producing S. marcescens in an outbreak at a public hospital in Cuiaba, MT, Brazil. METHODOLOGY: Ten isolates of S. marcenses were sequenced and antibiotic resistance profiles analyzed over 12 days. RESULTS: The isolates were clonal and multidrug resistant. Gentamycin and tigecycline had sensitivity in 90% and 80% isolates, respectively. Genomic analysis identified several genes that encode ß-lactamases, aminoglycoside-modifying enzymes, efflux pumps, and other virulence factors. CONCLUSIONS: Systematic surveillance is crucial in monitoring the evolution of S. marcescens genotypes, as it can lead to early detection and prevention of outbreaks.

Brazilian porcupinepox virus infection in a free-ranging neotropical porcupine in Mato Grosso, Brazil.

Although the novel Brazilian porcupinepox virus (BPoPV) can infect wild porcupines, its lethality and zoonotic potential are not well-established. In May 2021, a free-ranging neotropical porcupine (Coendou longicaudatus boliviensis) rescued from the natural savanna in the Brazilian Cerrado, Mato Grosso State, was presented with a lethal poxviral infection. Clinical signs and PCR detection of BPoPV supported the diagnosis. Poxviral lesions included erythema, exfoliative dermatitis, and erosions involving mainly the face, hindlimb, and vulva. Histologically, the lesions consisted of proliferative and necrotic dermatitis, intraepidermal and follicular pustules, and intracytoplasmic inclusion bodies in keratinocytes. Phylogenetic analysis revealed BPoPV strains closely related to other rodent-infecting poxviruses. This poxviral infection resulted in the death of a specimen of C. l. boliviensis; the effect on species conservation, and the potential of spillover into humans and other vertebrates remain unknown.

KPC-2-producing Pseudomonas aeruginosa isolated from wild animals in Brazil.

Pseudomonas aeruginosa an opportunistic pathogen that causes infections in hospitals and has high morbidity and mortality rates. In addition, it is a widely distributed environmental bacterium that can colonise a variety of habitats. Although wild animals do not have access to antibiotics, antibacterial resistance in these animals has increasingly been reported worldwide. Although the presence of Klebsiella pneumoniae carbapenemase (KPC) is uncommon in P. aeruginosa, it has been increasingly reported. This study examined KPC-2-producing P. aeruginosa in wild animals. A total of 27 P. aeruginosa isolates were obtained from clinical cases treated at the Microbiology Laboratory of the Veterinary Hospital of UFMT, Brazil. P. aeruginosa and blaKPC-2 carbapenemase resistance genes were identified using PCR. Antimicrobial susceptibility of KPC-producing P. aeruginosa was evaluated using the disk diffusion method. The blaKPC-2 gene was detected in 40.7% of the isolates (11/27). The rates of antimicrobial resistance and intermediate sensitivity were as follows: piperacillin/tazobactam (44.4%), imipenem (29.6%), meropenem (51.8%), amikacin (77.8%), cefepime (85.2%), and ciprofloxacin (70.4%). Twelve isolates were classified as Multidrug-resistant (MDR). This study presents the first report of P. aeruginosa with the blaKPC-2 gene in wild animals in Brazil, highlighting the importance of molecular research on resistance genes in P. aeruginosa from a One-Health perspective.

Molecular prevalence and factors associated with Ehrlichia canis infection in dogs from the North Pantanal wetland, Brazil.

Background and Aim: Canine monocytic ehrlichiosis is a vector-borne disease caused by the obligatory intracellular bacterium Ehrlichia canis, which is distributed across tropical and subtropical regions worldwide. Its prevalence within dog populations is high in municipalities located across the Pantanal biome, but it remains unknown in Barão de Melgaço, Mato Grosso, Brazil. This study aimed to determine the molecular prevalence and factors associated with E. canis infection in dogs domiciled in Barão de Melgaço. Materials and Methods: A cross-sectional study was carried out to investigate the prevalence of E. canis infection in 369 dogs from urban and rural areas in Barão de Melgaço, North Pantanal wetland, Brazil. Initially, the dogs were examined, and, through a questionnaire, the risk factors were investigated. Blood samples were subjected to DNA extraction and PCR was performed to estimate the prevalence of E. canis infection. Results: The molecular prevalence of E. canis infection in dogs was 42.5% and none of the studied variables were significantly associated with polymerase chain reaction (PCR) positivity (p > 0.05). Conclusion: The high molecular prevalence demonstrates an increased transmission of the agent across the city. This also indicates that attention needs to be paid to E. canis infection and control measures should be introduced to prevent its transmission. The demographic and clinical risk factors commonly associated with E. canis infection in this study were not associated with PCR positivity.

Molecular detection and genetic characterization of Ehrlichia canis and Ehrlichia sp. in neotropical primates from Brazil.

The Anaplasmataceae family includes obligate, arthropod-transmitted intracellular bacteria that can be zoonotic and potentially fatal. Studies focusing on the interaction between neotropical primates and the agents of this family are scarce. The present study aimed to identify agents of the Anaplasmataceae family in the whole blood of free-living and captive neotropical primates in the State of Mato Grosso, Central-West Brazil. Thirty-eight samples of six nonhuman primate (NHP) species were collected in seven municipalities and analysed through polymerase chain reaction (PCR), nucleotide sequencing, and phylogenetic analysis of the dsb, groEL, 16S rRNA, and gltA genes. DNA fragments similar to those of Ehrlichia canis were detected in Sapajus apella and Ehrlichia chaffeensis from Mico melanurus. The sequences generated in this study and homologous sequences retrieved from GenBank® were used for phylogenetic analyses to characterize the Ehrlichial agents detected in NHPs. The agents were then grouped into clades corresponding to different isolates from the NHP species. In addition, an Anaplasma sp. closely related to Anaplasma marginale was identified in two S. apella individuals. These findings shed light on the susceptibility of neotropical NHPs to Anaplasmataceae agents. These bacteria are known to be transmitted by ticks, which can also serve as possible sources of infection for other animals, including humans.

SARS-CoV-2 antibodies in dogs and cats in a highly infected area of Brazil during the pandemic.

SARS-CoV-2 was a worldwide threat during the COVID-19 pandemic, and the state of Mato Grosso had the second highest mortality rate in Brazil, with 427. 4 deaths/100,000 inhabitants. However, no large-scale study among dogs and cats in such highly infected areas of Brazil has so far been conducted. Accordingly, the present study reports on a serosurvey among dogs and cats in Cuiabá, capital of Mato Grosso from November 2020 to July 2021, where the human mortality rate was 605/100,000 at that time. Overall, 33/762 dogs (4.3%) and 4/182 cats (2.2%) were found to be seropositive for SARS-CoV-2 through ELISA, and 3/762 dogs (0.4%) and 3/182 cats (1.6%) were seropositive through the serum neutralization test. Cats presented higher seroprevalence with higher titers of neutralizing antibodies. Although N-protein based ELISA may be a good screening test, cross-reactivity with other canine coronaviruses may impair its diagnostic use among dogs.