Crithidia ricardoi sp. n. a new species of trypanosomatidae isolated from Culex saltanensis Dyar, 1928 (Diptera: Culicidae)

A flagellte trypanosomatid was isolated in culture from the digestive tract of the mosquito Culex saltanensis Dyar, 1928. It grows exuberantly in liver infusion in the form of epimastigotes and choanomastigotes typical of the genus Crithidia. The trypanosomatid was compared to C. deanei, C. fasciculata, C. luciliae, C. oncopelti and C. guilhermei. The techniques used for comparison were electron transmission microscopy, isoenzymes and kDNA restriction profiles. No endosymbionts were found at electron microscopy. Results for the biochemical methods employd indicate that the trypanosomatid isolated from C. saltanensis is a new species of Crithidia. The name C. ricardoi sp. n. is proposed for thes new species.

Survey of the irp2 gene among Yersinia pestis strains isolated during several plague outbreaks in northeast Brazil

The irp2 gene codes for a 190 kDa protein (HMWP2) synthesidez when highly pathogenic Yersinia are grown under conditions of iron starvation. In this work, the presence of irp2 in strains of Y. pestis isolated from different hosts during several plague outbreaks in the foci of Northeast Brazil wasstudied. For this purpose, 53 strains were spotted onto nylon filters and their DNA was hybridized with the A13 probe which is a 1 kb fragment of the irp2 coding sequence. All strains except two hybridized with the probe. However, when the initial stock culture of these two strains were analyzed, they both proved to bepositive with the A13 probe, indicating that the locus was lost after subculturein vitro but was always present in vivo. To examine the degree of conservation of the chromosomal fragment carrying irp2 among Brazilian strains, the hybridization profiles of 15 strains from different outbreaks, different hosts and different foci were compared. The hybridization profiles of these strains were all identical when their DNA was digested with either EcoRI, EcorRV or AvaII, indicatingthat the restriction sites surrounding the irp2 locus are very well conserved among Northeast Brazilian strains of Y. pestis. Altogether, these results suggest that the irp2 chromosomal region should be of prime importance for the bacteria during their multiplication in the host.

Intratypic differentiation of polioviruses isolated from suspected cases of poliomyelitis in Brazil during the period of 1990 to 1993

This study analyzed 3129 fecal samples derived from 1626 patients with sudden onset acute flaccid paralysis clinically compatible with poliomyelitis. The samples were collected in the period ranging from January 1990 to September 1993 in all regions of Brazil. Among the 1626 cases studied, 196 had isolation of poliovirus. Nevertheless, it was observed that some factors influenced the isolation rate and the intratypic characterization of these polioviruses. No cases of acute flaccid paralysis has been found to be etiologically related with wild polioviruses.

CHO expression of a novel human recombinant IgG1 anti-RhD antibody isolated by phage display.

Replacement of the hyperimmune anti-Rhesus (Rh) D immunoglobulin, currently used to prevent haemolytic disease of the newborn, by fully recombinant human anti-RhD antibodies would solve the current logistic problems associated with supply and demand. The combination of phage display repertoire cloning with precise selection procedures enables isolation of specific genes that can then be inserted into mammalian expression systems allowing production of large quantities of recombinant human proteins. With the aim of selecting high-affinity anti-RhD antibodies, two human Fab libraries were constructed from a hyperimmune donor. Use of a new phage panning procedure involving bromelin-treated red blood cells enabled the isolation of two high-affinity Fab-expressing phage clones. LD-6-3 and LD-6-33, specific for RhD. These showed a novel reaction pattern by recognizing the D variants D(III), D(IVa), D(IVb), D(Va), D(VI) types I and II. D(VII), Rh33 and DFR. Full-length immunoglobulin molecules were constructed by cloning the variable regions into expression vectors containing genomic DNA encoding the immunoglobulin constant regions. We describe the first, stable, suspension growth-adapted Chinese hamster ovary (CHO) cell line producing a high affinity recombinant human IgG1 anti-RhD antibody adapted to pilot-scale production. Evaluation of the Fc region of this recombinant antibody by either chemiluminescence or antibody-dependent cell cytotoxicity (ADCC) assays demonstrated macrophage activation and lysis of red blood cells by human lymphocytes. A consistent source of recombinant human anti-RhD immunoglobulin produced by CHO cells is expected to meet the stringent safety and regulatory requirements for prophylactic application.

Analysis of the clonal diversity of Staphylococcus aureus methicillin-resistant strains isolated at João Pessoa, state of Paraíba, Brazil

To investigate the clonal diversity of Staphylococcus aureus strains isolated at João Pessoa, State of Paraíba, Brazil, digested genomic DNA were studied by pulsed-field gel electrophoresis (PFGE) in nine methicillin-resistant strains (MRSA) and three methicillin-sensitive strains (MSSA), selected among 67 isolates based on their antimicrobial susceptibility and epidemiology. The isolates were obtained between April and November 1992 from the Hospital of the Federal University of Paraíba, located in João Pessoa. Two MRSA isolates from the Oswaldo Cruz Hospital, São Paulo, Brazil, including an epidemic strain previously detected from different hospitals at the country were used as control. Five different patterns, were demonstrated by MRSA isolated in João Pessoa and these patterns were described in several epidemiologically unrelated hospitals in São Paulo. Our results suggest the interstate dissemination of a MRSA clone in João Pessoa which is similar to that described in other cities of Brazil.

N-terminal amino acid sequences of the major outer membrane proteins from a Neisseria meningitidis group B strain isolated in Brazil

The four dominant outer membrane proteins (46, 38, 33 and 28 kDa) were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in a semi-purified preparation of vesicle membranes of a Neisseria meningitidis (N44/89, B:4:P1.15:P5.5,7) strain isolated in Brazil. The N-terminal amino acid sequence for the 46 kDa and 28 kDa proteins matched that reported by others for class 1 and 5 proteins respectively, whereas the sequence (25 amino acids) for the 38 kDa (class 3) protein was similar to class 1 meningococcal proteins. The sequence for the 33 kDa (class 4) was unique and not homologous to any known protein.

Characterization of Trypanosoma cruzi Strains Isolated from Chronic Chagasic Patients, Triatomines and Opossums Naturally Infected from the State of Rio Grande do Sul, Brazil

Thirty-five Trypanosoma cruzi strains were isolated from chronic chagasic patients, triatomines and opossums from different municipalities of the State of Rio Grande do Sul. Parasites were characterized by means of mice infectivity, enzyme electrophoresis and randomly amplified polymorphic DNA (RAPD) analysis. Twenty-nine strains were isolated from chagasic patients, 4 from triatomines (2 from Triatoma infestans and 2 from Panstrongylus megistus) and 2 from opossums Didelphis albiventris. Thirty-three T. cruzi strains were of low and 2 strains of high virulence in mice. Both virulent strains were isolated from P. megistus. Isoenzyme analysis of the strains showed 3 different zymodemes. Eleven strains isolated from chagasic patients and 2 from D. albiventris were Z2. Eighteen strains from patients and 2 from T. infestans were ZB and 2 T. cruzi strains isolated from P. megistus were Z1. RAPD profiles obtained with 4 random primers showed a high genetic heterogeneity of the T. cruzi strains. Zymodeme 2 and ZB strains were the more polymorphic. A band sharing analysis of the RAPD profiles of Z2 and ZB strains using 3 primers, showed a very low percentage of shared bands, 20% among 13 ZB strains and 14% among 13 Z2 strains. According to the isoenzyme results, 3 T. cruzi populations were present in State of Rio Grande do Sul. Zymodeme 2 and ZB strains were found infecting man (domiciliar transmission cycle) whereas Z1 strains were found infecting the sylvatic vector P. megistus

Analysis of Mycobacterium avium Complex Serovars Isolated from AIDS Patients from Southeast Brazil

The purpose of this study was to assess the distribution of Mycobacterium avium serovars isolated from AIDS patients in São Paulo and Rio de Janeiro. Ninety single site or multiple site isolates from 75 patients were examined. The most frequent serovars found were 8 (39.2%), 4 (21.4%) and 1 (10.7%). The frequency of mixed infections with serovar 8 or 4 was 37.8%. Among the 90 strains examined, M. intracellulare serovars (7 strains) and M. scrofulaceum (4 strains) were found in 11 isolates (12%) indicating that M. avium (88%) was the major opportunistic species in the M. avium complex isolates in Brazilian AIDS patients

Plasmodium falciparum: nitric oxide modulates heme speciation in isolated food vacuoles.

Nitric oxide (NO) and NO-derived reactive nitrogen species (RNS) are present in the food vacuole (FV) of Plasmodium falciparum trophozoites. The product of PFL1555w, a putative cytochrome b(5), localizes in the FV membrane, similar to what was previously observed for the product of PF13_0353, a putative cytochrome b(5) reductase. These two gene products may contribute to NO generation by denitrification chemistry from nitrate and/or nitrite present in the erythrocyte cytosol. The possible coordination of NO to heme species present in the food vacuole was probed by resonance Raman spectroscopy. The spectroscopic data revealed that in situ generated NO interacts with heme inside the intact FVs to form ferrous heme nitrosyl complexes that influence intra-vacuolar heme solubility. The formation of heme nitrosyl complexes within the FV is a previously unrecognized factor that could affect the equilibrium between soluble and crystallized heme within the FV in vivo.

Serotypes of Vibrio cholerae Non-O1 Isolated from Water Supplies for Human Consumption in Campeche, México and their Antibiotic Susceptibility Pattern

The presence of Vibrio cholerae non-O1 in water supplies for human consumption in the city of Campeche and rural locality of Bécal was investigated. V. cholerae non-O1 was detected in 5.9% of the samples obtained in deep pools of Campeche. Studies conducted in Bécal and neighbourhood of Morelos in Campeche indicated that collected samples harbored V. cholerae non-O1 in 31.5% and 8.7% respectively. There was a particular pattern of distribution of V. cholerae non-O1 serotypes among different studied regions. Accordingly, V. cholerae non-O1 serotype O14 predominated in the deep pools of Campeche and together with V. cholerae non-O1, O155 were preferentially founds in samples taken from intradomiciliary faucets in the neighbourhood of Morelos. Samples from Bécal predominantly presented the serotype O112. 60% and 53.8% of all studied strains of V. cholerae non-O1 proved to be resistant to ampicillin and carbenicillin. 3.1%, 7.7% and 6.2% presented resistant to doxycycline, trimethoprim-sulfamethoxazole and erythromycin respectively. The study showed the necessity of performing a strong epidemiologic surveillance for emergence and distribution of V. cholerae non-O1