An unknown Trypanosoma species was isolated from an axenic culture of intact skin from a domestic dog captured in Rio de Janeiro, Brazil, which was co-infected with Leishmania (Viannia) braziliensis. Giemsa-stained smears of cultures grown in different media revealed the presence of epimastigotes, trypomastigotes, spheromastigotes, transitional stages, and dividing forms (epimastigotes or spheromastigotes). The highest frequency of trypomastigotes was observed in RPMI (15.2%) and DMEM (9.2%) media containing 5% FCS, with a mean length of these forms of 43.0 and 36.0 mum, respectively. Molecular analysis by sequential application of PCR assays indicated that this trypanosome differs from Trypanosoma cruzi and T. rangeli when specific primers were applied. On the other hand, a PCR strategy targeted to the D7 domain of 24salpha rDNA, using primers D75/D76, amplified products of about 250 bp in that isolate (stock A-27), different from the amplification products obtained with T. cruzi and T. rangeli. This organism differs from T. cruzi mainly by the size of its trypomastigote forms and kinetoplasts and the absence of infectivity for macrophages and triatomine bugs. It is also morphologically distinct from salivarian trypanosomes reported in Brazil. Isoenzyme analysis at 8 loci demonstrated a very peculiar banding pattern clearly distinct from those of T. rangeli and T. cruzi. We conclude that this isolate is a new Trypanosoma species. The name T. caninum is suggested.

Promastigote forms of a trypanosomatid were isolated from the third and fourth ventricles of the midgut and from the hindgut of the phytophagous hemipteran Oncopeltus fasciatus. Some individuals had adhered to its anterior region, close to the flagellar pocket, or to the flagellum up to four rounded aflagellated forms known as straphangers cysts. Scanning electron microscopy revealed that the flagellated forms presented a twisted cell body and a long flagellum, and the cysts, smaller than the parental promastigote, had a nascent flagellum. Transmission electron microscopy showed that promastigotes were typical, while cystic forms were ovoid dense cells devoid of a cyst wall, but presenting a cell coat, a special subpellicular region limited by a membrane unit, and a condensed cytoplasm. The kinetoplast-DNA fibrils appeared as dense spots and the condensed chromatin was arranged in a labyrinthic structure. Desmosome-like structures, observed in the region of adhesion of the precystic forms to the parental promastigote, could explain how cysts remain attached to the mother cell during the encystation process. Release of membranes from the surface of promastigotes and cysts seems to be correlated with the condensation of the cytoplasm during encystment. Morphological and isozyme analyses indicated that this trypanosomatid belongs to the genus Leptomonas. The molecular karyotype of this isolate was compared with that of a strain of Leptomonas oncopelti obtained from Oncopeltus varicolor by contour-clamped homogeneous electric field (CHEF) electrophoresis and revealed similar DNA banding patterns between 2,200-825 Kb, but not in lower bands (825-225 Kb). This suggested that the isolate from O. fasciatus and that from O. varicolor were not identical. Based on our findings we are describing Leptomonas wallacei n. sp. for our isolate from O. fasciatus.

for the BaSICS investigators and the BRICNet members IMPORTANCE Slower intravenous fluid infusion rates could reduce the formation of tissue edema and organ dysfunction in critically ill patients; however, there are no data to support different infusion rates during fluid challenges for important outcomes such as mortality.OBJECTIVE To determine the effect of a slower infusion rate vs control infusion rate on 90-day survival in patients in the intensive care unit (ICU). DESIGN, SETTING, AND PARTICIPANTS Unblinded randomized factorial clinical trial in 75 ICUs in Brazil, involving 11 052 patients requiring at least 1 fluid challenge and with 1 risk factor for worse outcomes were randomized from May 29, 2017, to March 2, 2020. Follow-up was concluded on October 29, 2020. Patients were randomized to 2 different infusion rates (reported in this article) and 2 different fluid types (balanced fluids or saline, reported separately).INTERVENTIONS Patients were randomized to receive fluid challenges at 2 different infusion rates; 5538 to the slower rate (333 mL/h) and 5514 to the control group (999 mL/h). Patients were also randomized to receive balanced solution or 0.9% saline using a factorial design. MAIN OUTCOMES AND MEASURESThe primary end point was 90-day survival.RESULTS Of all randomized patients, 10 520 (95.2%) were analyzed (mean age, 61.1 years [SD, 17.0 years]; 44.2% were women) after excluding duplicates and consent withdrawals. Patients assigned to the slower rate received a mean of 1162 mL on the first day vs 1252 mL for the control group. By day 90, 1406 of 5276 patients (26.6%) in the slower rate group had died vs 1414 of 5244 (27.0%) in the control group (adjusted hazard ratio, 1.03; 95% CI, 0.96-1.11; P = .46). There was no significant interaction between fluid type and infusion rate (P = .98).CONCLUSIONS AND RELEVANCE Among patients in the intensive care unit requiring fluid challenges, infusing at a slower rate compared with a faster rate did not reduce 90-day mortality. These findings do not support the use of a slower infusion rate.

A method to purify trypanosomastigotes of some strains of Trypanosoma cruzi (Y, CL, FL, F, "Berenice", "Colombiana" and "São Felipe") from mouse blood by using DEAE-cellulose columns was standardized. This procedure is a modification of the Lanham & Godfrey methods and differs in some aspects from others described to purify T. cruzi bloodstream trypomastigotes, mainly by avoidance of prior purifications of parasites. By this method, the broad trypomastigotes were mainly isolated, accounting for higher recoveries obtained with strains having higher percentages of these forms: processing of infected blood from irradiated mice could be advantageous by increasing the recovery of parasites (percentage and/or total number) and elution of more slender trypomastigotes. Trypomastigotes purified by this method presented normal morphology and motility, remained infective to triatomine bugs and mice, showing in the latter prepatent periods and courses parasitemia similar to those of control parasites, and also reproducing the polymorphism pattern of each strain. Their virulence and pathogenicity also remained considerably preserved, the latter property being evaluated by LD 50 tests, mortality rates and mean survival time of inoculated mice. Moreover, these parasites presented positive, clear and peripheral immunofluorescence reaction at titres similar to those of control organisms, thus suggesting important preservation of their surface antigens.
Usando colunas de DEAE-cellulose foi padronizado um método para purificação de tripomastigotas de várias cepas de Trypanosoma cruzi (Y, CL, FL, F, "Berenice", "Columbiana" e "São Felipe") a partir do sangue de camundongos. Este método é uma modificação daqueles descritos por Lanham & Godfrey e difere em vários aspectos de outros descritos para purificar as formas sanguíneas deste parasita, particularmente na dispensa de pré-purificações. Por ele foram isolados principalmente os tripomastigotas largos, oque certamente justifica as maiores percentagens de recuperação obtidas com cepas em que predominavam estas formas; por outro lado, o processamento de sangue infectado de camundongos irradiados podia ser vantajoso pela recuperação de maior número e percentagem de parasitas e eluição de mais formas finas. Os tripomastigotas purificados por este método apresentavam morfologia e motilidade normais e continuavam infectantes para barbeiros e camundongos. Nestes últimos apresentaram período pré-patente e curvas parasitêmicas semelhantes aos dos parasitas controles, além de reproduzirem o padrão de polimorfismo típico da cepa. Sua virulência e patogenicidade também manteve-se consideravelmente preservada, sendo que esta última propriedade foi avaliada por testes de DL 50, taxas de mortalidade e tempo médio de sobrevida de camundongos inoculados. Além do masi, os tripomastigotas purificados apresentaram reação de imunofluorescência positiva, nítida e periférica, cujos títulos eram comparáveis aos dos parasitas controles, assim sugerindo uma importante preservação de seus antígenos de supe...

We report the finding, the isolation by hemoculture, and the characterization of Trypanosoma rangeli stocks from two chronic Chagas' disease patients who received ambulatory care at the Evandro Chagas Clinical Research Institute (IPEC, FIOCRUZ). Both patients proceeded from Bahia State (Brazil). One of them presented the cardiac form of the disease and the other indeterminate symptomalogy. Giemsa-stained smears of the hemocultures from these patients evidenced that they were coinfected with T. rangeli and Trypanosoma cruzi, with predominance of the former species. These isolates could only be successfully grown in Novy-MacNeal-Nicolle + liver infusion-tryptose supplemented with 20-30% fetal calf serum. After 6 months of serial maintenance, rich and apparently pure cultures of T. rangeli were obtained. Both stocks were analyzed with different approaches and compared with two T. cruzi isolates also from chagasic patients under care at IPEC, besides T. rangeli and T. cruzi reference strains. All stocks were characterized by morphology, biometry, electrophoresis of isoenzymes, and products of kDNA minicircle amplified by polymerase chain reaction. The identification of T. rangeli was largely confirmed by all techniques. Taken together, these data represent the third report on T. rangeli in human hosts in Brazil.