Case Report: SARS-CoV-2 Gamma Isolation From Placenta of a Miscarriage in Midwest, Brazil.

The present study investigated a SARS-CoV-2 infection in placenta and fetal samples from an early pregnancy miscarriage in Midwest Brazil. The Gamma variant was isolated and fully sequenced from the placenta sample, but not from fetal samples. Our findings highlight potential adverse perinatal outcomes caused by SARS-CoV-2 Gamma infection during pregnancy.

Molecular Survey of Bartonella Species in Shelter Cats in Rio De Janeiro: Clinical, Hematological, and Risk Factors.

The present study aimed to detect Bartonella DNA in cats belonging to shelters, and to evaluate risk factors, clinical signs, and hematological abnormalities associated with infection. Complete blood counts and screening for the presence of Bartonella DNA were performed on cats' ethylenediamine tetraacetic acid anticoagulant-blood samples. Eighty-three cats (39.9%) were positive for Bartonella species. Bartonella DNA was also detected in fleas and in the blood of cats infested by positive flea. Cats that had not been sterilized, had outdoor access, had histories of fights, and had concurrent flea infestation were more likely to be infected by Bartonella species (P < 0.05). Age and sex were not associated with infection. Fifty-one (38.6%) symptomatic cats were positive to Bartonella species (P > 0.05). Clinical conditions most commonly observed were signs of respiratory abnormality and Sporothrix species coinfection (P > 0.05). Regarding hematological changes, eosinophilia was associated with infection (P < 0.05). A high frequency of Bartonella species infection was found in shelter cats and highlights the importance of adequate flea-control programs to prevent infection in cats and consequently in adopters and other animals.

Molecular identification of Bartonella henselae in a seronegative cat scratch disease patient with AIDS in Rio de Janeiro, Brazil.

Bartonella henselae is associated with a wide spectrum of clinical manifestations, including cat scratch disease, endocarditis and meningoencephalitis, in immunocompetent and immunocompromised patients. We report the first molecularly confirmed case of B. henselae infection in an AIDS patient in state of Rio de Janeiro, Brazil. Although DNA sequence of B. henselae has been detected by polymerase chain reaction in a lymph node biopsy, acute and convalescent sera were nonreactive.

MOLECULAR IDENTIFICATION OF Bartonella henselae IN A SERONEGATIVE CAT SCRATCH DISEASE PATIENT WITH AIDS IN RIO DE JANEIRO, BRAZIL / Identificação molecular de Bartonella henselae em paciente com SIDA soronegativo para doença da arranhadura do gato no Rio de Janeiro, Brasil

Bartonella henselae is associated with a wide spectrum of clinical manifestations, including cat scratch disease, endocarditis and meningoencephalitis, in immunocompetent and immunocompromised patients. We report the first molecularly confirmed case of B. henselae infection in an AIDS patient in state of Rio de Janeiro, Brazil. Although DNA sequence of B. henselae has been detected by polymerase chain reaction in a lymph node biopsy, acute and convalescent sera were nonreactive.

Bartonella henselae está associada a um amplo espectro de manifestações clínicas, incluindo a doença da arranhadura de gato, endocardite, e meningoencefalite, em pacientes imunocompetentes e imunocomprometidos. Relatamos o primeiro caso confirmado por método molecular de B. henselae em um paciente com SIDA no estado do Rio de Janeiro, Brasil. Apesar da sequência de DNA de B. henselae ser detectada pela reação em cadeia da polimerase em uma biópsia do linfonodo, soros das fases aguda e convalescente foram não reativos.

Detection of serum antibodies against Bartonella species in cats with sporotrichosis from Rio de Janeiro, Brazil.

Cat scratch disease is a zoonosis caused by Bartonella species, transmitted to humans through scratches or bites from infected cats and via direct contact with infected feces. Sporotrichosis, caused by the fungal complex Sporothrix, is transmitted by traumatic inoculation of the fungus. Cats are important in zoonotic transmission. Serum samples from 112 domestic cats with sporotrichosis and 77 samples from healthy cats were analyzed by indirect immunofluorescence assay (IFA), using the commercial kit Bartonella henselae IFA IgG (Bion). The presence of antibodies against feline leukemia virus (FeLV) and of feline immunodeficiency virus (FIV) core antigens was detected using the commercial kit Snap Combo FIV-FeLV (Idexx). The group of animals with sporotrichosis contained 93 males with a median age of 22 months, eight (7.1%) of which were positive for FIV and 15 (13.4%) for FeLV. The group of animals without sporotrichosis contained 36 males with a median age 48 months, 10 (13.0%) of which were positive for FIV and eight (10.4%) for FeLV. Of the 112 cats with sporotrichosis and 77 cats without mycosis, 72 (64.3%) and 35 (45.5%), respectively, were IFA reactive. No association was found between age, sex, FIV/FeLV and the presence of antibodies to Bartonella species. The results suggest that the study population can be considered a potential source of zoonotic infection for both diseases.

Coxiella burnetii, the agent of Q fever in Brazil: its hidden role in seronegative arthritis and the importance of molecular diagnosis based on the repetitive element IS1111 associated with the transposase gene.

Coxiella burnetii is the agent of Q fever , an emergent worldwide zoonosis of wide clinical spectrum. Although C. burnetii infection is typically associated with acute infection, atypical pneumonia and flu-like symptoms, endocarditis, osteoarticular manifestations and severe disease are possible, especially when the patient has a suppressed immune system; however, these severe complications are typically neglected. This study reports the sequencing of the repetitive element IS1111 of the transposase gene of C. burnetii from blood and bronchoalveolar lavage (BAL) samples from a patient with severe pneumonia following methotrexate therapy, resulting in the molecular diagnosis of Q fever in a patient who had been diagnosed with active seronegative polyarthritis two years earlier. To the best of our knowledge, this represents the first documented case of the isolation of C. burnetii DNA from a BAL sample.

Eschar-associated spotted fever rickettsiosis, Bahia, Brazil.

In Brazil, Brazilian spotted fever was once considered the only tick-borne rickettsial disease. We report eschar-associated rickettsial disease that occurred after a tick bite. The etiologic agent is most related to Rickettsia parkeri, R. africae, and R. sibirica and probably widely distributed from Sao Paulo to Bahia in the Atlantic Forest.

Bartonella spp. infection in HIV positive individuals, their pets and ectoparasites in Rio de Janeiro, Brazil: serological and molecular study.

BACKGROUND: Bartonella is the agent of cat-scratch disease, but is also responsible for more severe conditions such as retinitis, meningoencephalitis, endocarditis and bacillary angiomatosis. Its seroprevalence is unknown in Brazil. METHODS: Patients in an AIDS clinic, asymptomatic at the time of the study, were enrolled prospectively. They answered a structured questionnaire and had blood taken for serological and molecular assays. Cat breeder's pets were tested serologically and collected ectoparasites were tested by molecular biology techniques. Blood donors, paired by age and sex, were tested for Bartonella IgG antibodies. RESULTS: 125 HIV positive patients with a median age of 34 were studied; 61 were male and 75% were on HAART. Mean most recent CD4 count was 351-500 cells/mm(3). A high rate of contact with ticks, fleas and lice was observed. Bartonella IgG seroreactivity rate was 38.4% in HIV positive individuals and breeding cats was closely associated with infection (OR 3.6, CI 1.1-11.9, p<0.05). No difference was found between the sexes. Titers were 1:32 in 39 patients, 1:64 in seven, 1:128 in one and 1:256 in one. In the control group, IgG seroreactivity to Bartonella spp. was 34%, and female sex was correlated to seropositivity. Fourteen of 61 (23%) males vs 29/64 (45.3%) females were seroreactive to Bartonella (OR 2.8, CI 1.2-6.5, p<0.01). Titers were 1:32 in 29 patients, 1:64 in ten and 1:128 in four. CONCLUSIONS: Bartonella spp. seroprevalence is high in HIV positive and in blood donors in Rio de Janeiro. This may be of public health relevance.

First identification of natural infection of Rickettsia rickettsii in the Rhipicephalus sanguineus tick, in the State of Rio de Janeiro / Primeira identificação de infecção natural por Rickettsia rickettsii no carrapato Rhipicephalus sanguineus no Rio de Janeiro

The Brazilian Spotted Fever (BSF) is a zoonotic disease caused by Rickettsia rickettsii and transmitted by ticks of the genus Amblyomma, more frequently, Amblyomma cajennense. The aim of this paper was to report the first molecular detection of R. rickettsii on R. sanguineus naturally infected in Rio de Janeiro, Brazil. Ticks were collected from dogs in a rural region of Resende municipality, Rio de Janeiro State, Brazil (22º30'9.46"S, 44º42'44.29"WO), where occurred five human cases of BSF in 2006. The ticks were identified under a stereoscopic microscope and separated in pools by stages, species and sex. DNA extraction was carried out using QIAamp DNA Mini Kit (QIAGEN®). The DNA was submitted to PCR amplification using 04 set of primers: Rr190.70p/Rr190.602n (OmpA, 532bp), BG1-21/BG2-20 (OmpB, 650bp), Tz15/Tz16 (17 kDa protein-encoding gene, 246bp) and RpCS.877p/RpCS.1258n (gltA, 381bp). PCR products were separated by electrophoresis on 1 percent agarose gels and visualized under ultraviolet light with ethidium bromide. PCR products of the expected sizes were purified by QIAquick® and sequenced by ABI PRISM®. The generated nucleotide sequences were edited with using Bioedit® software and compared with the corresponding homologous sequences available through GenBank, using Discontiguous Mega Blast (http://www.ncbi.nlm.nih.gov). It was confirmed R. rickettsii by sequencing of the material (GenBank FJ356230). The molecular characterization of R. rickettsii in the tick R. sanguineus emphasizes the role of dogs as carriers of ticks from the environment to home. Moreover, this result suggests that there is a considerable chance for active participation of R. sanguineus as one of tick species in the transmission of R. ricketsii to human being in the Brazilian territory.

A Febre Maculosa Brasileira (FMB) é uma zoonose causada por Rickettsia rickettsii e transmitida por carrapatos do gênero Amblyomma, mais freqüentemente pela espécie Amblyomma cajennense. Este trabalho tem como objetivo relatar a primeira detecção molecular de R. rickettsii em Rhipicephalus sanguineus naturalmente infectado no Rio de Janeiro, Brasil. Carrapatos foram coletados de cães, procedentes de uma região rural do município de Resende, estado do Rio de Janeiro, Brasil (22º30'9.46"S, 44º42'44.29"WO), onde ocorreram cinco casos humanos de FMB em 2006. Todos os carrapatos foram identificados segundo chave dicotômica, utilizando-se lupa estereoscópica e separados de acordo com estágio, espécie e sexo. Para a extração de DNA utilizou-se o kit comercial QIAamp DNA (QIAGEN ®). O DNA foi submetido à técnica de PCR utilizando 04 conjuntos de iniciadores para a amplificação dos genes: Rr190.70p/Rr190.602n (OmpA, 532bp), BG1-21/BG2-20 (OmpB, 650bp), Tz15/Tz16 (17 kDa gene que codifica a proteína, 246bp) e RPCs .877p/RpCS.1258n (gltA, 381bp). Os produtos da PCR foram separados por eletroforese em gel agarose 1 por cento corados com brometo de etídio e visualizados sob luz ultravioleta e, aqueles que apresentaram bandas amplificadas foram purificados utilizando-se o kit comercial QIAquick ® e seqüenciados pelo ABI PRISM®. As seqüências nucleotídicas foram geradas usando Bioedit®, editado em software e comparados os correspondentes homólogos com as sequências disponíveis através GenBank, utilizando Discontiguous Mega Blast (http://www.ncbi.nlm.nih.gov). Confirmou-se R. rickettsii (GenBank FJ356230) no seqüenciamento de apenas um espécime, adulto de carrapato R. sanguineus. A caracterização molecular de R. rickettsii em exemplar de carrapato R. sanguineus confirma que esta espécie pode ter importante papel na transmissão de R. rickettsii para humanos no território brasileiro.

In vitro activity evaluation of Parkia pendula seed lectin against human cytomegalovirus and herpes virus 6.

Human cytomegalovirus (HCMV) in vitro infectivity was inhibited by Parkia pendula seed lectin (PpeL) in contrast to human herpes virus 6 (HHV-6) which was not affected. The antiviral activity was detected for HCMV in human embryo lung (HEL) cells using a microtechnique in culture plates. The assay showed a reduction of cellular infectivity from approximately 95%, at a concentration of 150microg/mL with minimal cytotoxicity (25%). Also, a reduction of 75% was observed in HEL cells at a concentration of 75microg/mL without toxic effect. The reduction on infectivity was observed even after virus pre-adsorption to cells suggesting that this action should occur after virus penetration, in the intracellular replication phase. MT4 lymphocytes and cord blood mononuclear cells (CBMC) were used to evaluate the lectin effect on HHV-6 following the same technique. Lectin concentrations with few or no toxic effects on lymphocytes did not show inhibitory action of HHV-6 cytopathic effect. The results obtained with PpeL demonstrate that it may have an impact in the design of pharmacological strategies to infection of cytomegalovirus.

Cloning, expression, and purification of recombinant bovine rotavirus hemagglutinin, VP8*, in Escherichia coli.

Rotavirus VP8* subunit is the minor trypsin cleavage product of the spike protein VP4, which is the major determinant of the viral infectivity and neutralization. To study the structure-function relationship of this fragment and to obtain type-specific reagents, substantial amounts of this protein are needed. Thus, full-length VP8* cDNA, including the entire trypsin cleavage-encoding region in gene 4, was synthesized and amplified by RT-PCR from total RNA purified from bovine rotavirus strain C486 propagated in MA104 cell culture. The extended VP8* cDNA (VP8ext) was cloned into the pGEM-T Easy plasmid and subcloned into the Escherichia coli expression plasmid pET28a(+). The correspondent 30 kDa protein was overexpressed in E. coli BL21(DE3)pLysS cells under the control of the T7 promoter. The identity and the antigenicity of VP8ext were confirmed on Western blots using anti-His and anti-rotavirus antibodies. Immobilized Ni-ion affinity chromatography was used to purify the expressed protein resulting in a yield of 4 mg of VP8ext per liter of induced E. coli culture. Our results indicate that VP8ext maintained its native antigenicity and specificity, providing a good source of antigen for the production of P type-specific immune reagents. Detailed structural analysis of pure recombinant VP8 subunit should allow a better understanding of its role in cell attachment and rotavirus tropism. Application of similar procedure to distinct rotavirus P serotypes should provide valuable P serotype-specific immune reagents for rotavirus diagnostics and epidemiologic surveys.