On the current existence of a coccidial line of development in the malaria parasites: a theory

Most opinion favors the origin of the malaria parasites from a coccidial ancestor. It is assumed that whatever the process through which the coccidia differentiated into a Plasmodium this phenomenon very probably occured millions of year ago, and during that differentiation process the original coccidia vanished. Therefore it has never repeated. At the light of some experiments the existence, at the present time, of a coccidial cycle of development in the malaria parasites, is proposed. The conection routes and mechanisms through which the malaria parasite changes to a coccidial life, and the routes in reverse are exposed. Transmission of the malaria-coccidial forms is suggested.

Human B19 parvovirus infetion: an example of multiple pathogenic effects determined by differences in host susceptibility

B19 infection offers some general lessons about human viruses and their possible effects on the human host, as follows: (1) Ubiquitous apparently benign viruses may have severe effects on a compromissed host. The virus may be invariable but the host can have diverse susceptibilities. (2) B19 and some other human viruses (through for none is the evidence so clear as for B19) have narrowly targetted effects. The host cell of B19 is a specialised progenitor of mature red cells: impairment of the function of this cell by B19 may cause profound anaemia. (3) The 'normal'host response to B19 may also cause disease, though this is slef limiting. (4) The effects of malfunction of the virus'target cell are exacerbated when the immune response is impaired by congenital or acquired immunodeficiency, immunosupressive therapy or, in the case of the fetus, developmental immaturity that allows the virus to persist.

Pharmacokinetic profile of two different pharmaceutical forms of theophylline (a slow release tablet and a syrup) after multiple dose administration to healthy human volunteers

Due to the narrow therapeutic range of theophyline, plasma concentrations of this drug are monitored in patients undergoing chronic therapy. Slow-release preparations avoid the fluctuations in plasma levels and improve patient compliance. In this study, we have compared the pharmacokinetic profiles of a theophylline slow-release tablet and a syrup form, when administered in multiple doses to healthy adult volunteers. The classification based upon releasing patterns is confirmed.

Immunogenicity of multiple antigen peptides containing Plasmodium vivax CS epitopes in BALB/c mice

Multiple antigen peptide systems (MAPs) allow the incorporation of various epitopes in to a single synthetic peptide immunogen. We have characterized the immune response of BALB/c mice to a series of MAPs assembled with different B and T cell epitopes derived from the Plasmodium vivax circumsporozoite (CS) protein. A B-cell epitope from the central repeat domain and two T-cell epitopes from the amino and carboxyl flanking regions were used to assembled eight different MAPs. An additional universal T cell epitope (ptt-30) from tetanus toxin protein was included. Immunogenicity in terms of antibody responses and in vitro T lymphocyte proliferation was evaluated. MAPs containing B and T cell epitopes induced high titers of anti-peptides antibodies, which recognized the native protein on sporozoites as determined by IFAT. The antibody specificity was also determined by a competitive inhibition assay with different MAPs. A MAP containing the B cell epitope (p11) and the universal epitope ptt-30 together with another composed of p11 and the promiscuous T cell epitope (p25) proved to be the most immunogenic. The strong antibody response and specificity for the cognate protein indicates that further studies designed to assess the potential of these proteins as human malaria vaccine candidates are warranted.

Establishment and Characterization of a Cell Line from the Mosquito Anopheles albimanus (Diptera: Culicidae)

A new cell line designated LSB-AA695BB, was established from embryos of the mosquito Anopheles albimanus. The primary culture was initiated in April, 1995, and the first passage was made 48 days later. Serial subcultures of the cells have been carried through 90 passages from Abril 1995 to February 1996. The cells were grown at 28°C in MK/VP12 medium, supplemented with 20% fetal bovine serum; the pH tolerance ranged between 6.8 to 7.0. The cells have also been adapted to MM/VP12 medium under the same pH, temperature and serum concentration. The majority of the cells were a fibroblast-type. Isozyme characterization showed a pattern similar to that of An. albimanus pupae and adults but distinct from Ae. taeniorhynchus and Ae. albopictus (C6/36) mosquito cell lines. The culture was shown to be free of mycoplasma, bacteria and fungi. Microsporidia contamination of transovarial transmission was controlled with 6.0 mg/ml of albendazole

Establishment and characterization of a new continuous cell line from Lutzomyia longipalpis (Diptera: Psychodidae) and its susceptibility to infections with arboviruses and Leishmania chagasi

Embryonic tissue explants of the sand fly Lutzomyia longipalpis (Lutz & Neiva 1912) the main vector of Leishmania chagasi (Cunha and Chagas), were used to obtain a continuous cell line (Lulo). The tissues were seeded in MM/VP12 medium and these were incubated at 28ºC. The first subculture was obtained 45 days after explanting and 96 passages have been made to date. Lulo is composed of epithelioid cells, showed a 0.04 generations/hour exponential growth rate and population doubling time at 24.7 h. The cell line isoenzymatic profiles were determined by using PGI, PGM, MPI and 6-PGDH systems, coinciding with patterns obtained from the same species and colony's pupae and adults. The species karyotype characteristics were recognized (2n = 8), in which pair 1 is subtelocentric and pairs 2, 3 and 4 are metacentric. Lulo was free from bacterial, fungal, mycoplasmic and viral infection. Susceptibility to five arbovirus was determined, the same as Lulo interaction with Leishmania promastigotes.

Multiple isolates from Aids patients: aspects of an analysis by a genotypic marker and antimicrobial susceptibilities variations

Twenty-one Mycobacterium avium multisolates, from ten human immunodeficiency virus-infected patients, were typed by restriction fragment length polymorphism using as marker the IS1245 and characterized by minimum inhibitory concentration for nine different antibiotics. Two out of four patients harboring multisolates with different fingerprint profile, were therefore considered as having a polyclonal infection, since their isolates were taken from sterile site. This result confirms that polyclonal infection caused by M. avium occurs with a nonnegligenciable frequency. Analyzing the multisolates susceptibility profile of each patient it was observed that most of them were infected with strains having appreciably different antimicrobial susceptibility patterns, no matter what the genotypic pattern of the strains was. These results have strong implication for the treatment of the patients.

Hepatitis C prevalence and risk factors in hemodialysis patients in Central Brazil: a survey by polymerase chain reaction and serological methods

An hemodialysis population in Central Brazil was screened by polymerase chain reaction (PCR) and serological methods to assess the prevalence of hepatitis C virus (HCV) infection and to investigate associated risk factors. All hemodialysis patients (n=428) were interviewed in eight dialysis units in Goiânia city. Blood samples were collected and serum samples screened for anti-HCV antibodies by an enzyme-linked immunosorbent assay (ELISA). Positive samples were retested for confirmation with a line immunoassay (LIA). All samples were also tested for HCV RNA by the PCR. An overall prevalence of 46.7% (CI 95%: 42-51.5) was found, ranging from 20.7% (CI 95%: 8.8-38.1) to 90.4% (CI 95%: 79.9-96.4) depending on the dialysis unit. Of the 428 patients, 185 were found to be seropositive by ELISA, and 167 were confirmed positive by LIA, resulting in an anti-HCV prevalence of 39%. A total of 131 patients were HCV RNA-positive. HCV viremia was present in 63.5% of the anti-HCV-positive patients and in 10.3% of the anti-HCV-negative patients. Univariate analysis of risk factors showed that the number of previous blood transfusions, transfusion of blood before mandatory screening for anti-HCV, length of time on hemodialysis, and treatment in multiple units were associated with HCV positivity. However, multivariate analysis revealed that blood transfusion before screening for anti-HCV and length of time on hemodialysis were significantly associated with HCV infection in this population. These data suggest that nosocomial transmission may play a role in the spread of HCV in the dialysis units studied. In addition to anti-HCV screening, HCV RNA detection is necessary for the diagnosis of HCV infection in hemodialysis patients.

A new continuous cell line from the mosquito Psorophora confinnis (Diptera: Culicidae) and its susceptibility to infections with some arboviruses

A new cell line, PC-0199-BR, was established from embryonated eggs of the mosquito Psorophora confinnis. To date (September 2000) it has had 62 continuous passages. This is the first report of a cell line of mosquitoes belonging to the genus Psorophora. Cell growth initially was achieved in the MM/VP12 medium, supplemented with 20% fetal bovine serum; however, the subcultures were later adapted to Grace's medium with 10% fetal bovine serum. Cell morphology in the primary cultures was heterogeneous; but later in the established cell line, the predominant cell type was epithelioid. Cultured cells were predominantly diploid (2n=6); however, chromosome abnormalities were observed in a small proportion of the cells in later passages. C and G band patterns were also determined in the karyotype. The cell line isozyme profiles coincided with pupae and adult samples of the species taken from the same colony. A preliminary arbovirus susceptibility study for the cell line was undertaken. No evidence was observed of contamination of the cell line with bacteria, fungi or mycoplasma.

Morphological investigation of Toxoplasma gondii in vivo by a multiple beam interference microscope

A recently developed technique, namely multiple beam interference microscopy, has been applied to investigate the morphology of the parasite Toxoplasma gondii for the first time. The interference pattern obtained from the multiple internal reflection of a T. gondii, sandwiched between a glass plate and a cover plate, was focused on the objective of a conventional microscope. Because of the enhance contrast, several details of sub cellular structure and separating compartments are clearly visible. Details reveal the presence of a nucleus, lipid body, dense granule, rhoptry and amylopectin. The wall thickness of the membrane of the lipid body and the amylopectin is of the order of 0.02 µm and can be clearly distinguished with the help of the present technique. The same parasite has also been examined with the help of atomic force microscopy, and because of its thick membrane, the inner structural details were not observed at all. Sub cellular details of T. gondii observed with the present technique have been reported earlier only by low amplification transmission electron microscopy and not by any optical microscopic technique.

Laboratory diagnosis of Schistosomiasis in areas of low transmission: a review of a line of research

After 57 years of successful control of schistosomiasis in Venezuela, the prevalence and intensity of infection have declined. Approximately 80% of the individuals eliminate less than 100 eggs/g of stools, therefore morbidity is mild and the majority are asymptomatic. The sensitivity of Kato-Katz decreases to approximately 60%. Available serological methods for the detection of circulating antigens only reach a 70% of sensitivity. Tests based on the detection of antibodies by immunoenzymatic assays have been improved. The circumoval precipitine test has shown a high sensitivity (97%), specificity (100%), and correlation with oviposition, being considered the best confirmatory diagnostic test. Additionally to the classical immunoenzymatic assays, the development of the alkaline phosphatase immunoassay, allowed to reach a 100% specificity with an 89% sensitivity. Recently, we have developed a modified ELISA in which the soluble egg antigen is treated with sodium metaperiodate (SMP-ELISA) in order to eliminate the glycosilated epitopes responsible for the false positive reactions. The specificity and sensitivity reaches 97% and 99%, respectively. Synthetic peptides from the excretory-secretory enzymes, cathepsin B (Sm31) legumain (Sm32) and cathepsin D (Sm45), have been synthesized. The combination of two peptides derived from the Sm31 have been evaluated, reaching a sensitivity of 96% when analyzed independently and with a 100% specificity. Antibodies raised in rabbits against peptides derived from the Sm31 and Sm32 are currently evaluated in two different antigen-capture-based assays. The development of a simple, cheap and reliable test that correlates with parasite activity is a major goal.

Evaluation of indirect susceptibility testing of Mycobacterium tuberculosis to the first- and second-line, and alternative drugs by the newer MB/BacT system

In order to evaluate the Organon Teknika MB/BacT system used for testing indirect susceptibility to the alternative drugs ofloxacin (OFLO), amikacin (AMI), and rifabutin (RIF), and to the usual drugs of standard treatment regimes such as rifampin (RMP), isoniazid (INH), pyrazinamide (PZA), streptomycin (SM), ethambutol (EMB), and ethionamide (ETH), cultures of clinical specimens from 117 patients with pulmonary tuberculosis under multidrug-resistant investigation, admitted sequentially for examination from 2001 to 2002, were studied. Fifty of the Mycobacterium tuberculosis cultures were inoculated into the gold-standard BACTEC 460 TB (Becton Dickinson) for studying resistance to AMI, RIF, and OFLO, and the remaining 67 were inoculated into Lowenstein Jensen (LJ) medium (the gold standard currently used in Brazil) for studying resistance to RMP, INH, PZA, SM, EMB, and ETH. We observed 100% sensitivity for AMI (80.8-100), RIF (80.8-100), and OFLO (78.1-100); and 100% specificity for AMI (85.4-100), RIF (85.4-100), and OFLO (86.7-100) compared to the BACTEC system. Comparing the results obtained in LJ we observed 100% sensitivity for RMP (80-100), followed by INH - 95% (81.8-99.1), EMB - 94.7% (71.9-99.7), and 100% specificity for all drugs tested except for PZA - 98.3 (89.5-99.9) at 95% confidence interval. The results showed a high level of accuracy and demonstrated that the fully automated, non-radiometric MB/BacT system is indicated for routine use in susceptibility testing in public health laboratories.