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L Oskam

Publications and source records attributed to L Oskam.

At least 37 records · Page 2Linked to original sources
PubMed1997

Visceral leishmaniasis from Bal'a, Palestine, caused by Leishmania donovani s.1. identified through polymerase chain reaction and restriction fragment length polymorphism analysis.

A 5-year old female from Bal'a, Tulkarm area, Palestine, was admitted with an 8 month history of fever, excessive night sweating, abdominal distension and enlargement, weight loss and sever anorexia. She was investigated elsewhere without reaching specific diagnosis. On admission, the history and symptoms were compatible with visceral leishmaniasis and bone marrow aspirate was positive for Leishmania amastigotes. The serum titer, using IFAT, was 1:640 for L. infantum and 1:320 for L. major promastigotes. When bone marrow material was also subjected to PCR followed by RFLP enzyme analysis, three fragments of the PCR product of the parasite present were obtained: two fragments of 260 bp and one fragment of 80 bp, identical with the pattern obtained with L. donovani. The patient received sodium stibogluconate, 200 mg IM for 30 days. Six months after treatment, the spleen was 2 cm below the costal margin, the liver was not palpable and she gained 1 kg. This case alerts general practitioners, pediatricians and health authorities to the presence of visceral leishmaniasis in Palestine and to the possibility of the disease being encountered in Jordan.

Animals↗
PubMed1997

Use of PCR on lymph-node sample as test of cure of visceral leishmaniasis.

When the polymerase chain reaction (PCR) was used to test lymph-node aspirates from 35 patients from eastern Sudan, who had had visceral leishmaniasis but were believed cured, leishmanial DNA was detected in samples from 14 of the patients. There were no significant differences between the PCR-positives and -negatives in terms of age, sex, spleen size, malaria status or presence of anti-Leishmania antibodies. However, PCR was more often positive in the patients who tested negative by the leishmanin skin test (LST) than in those who gave positive skin tests. Moreover, patients with a positive PCR and a negative LST converted more often to LST positivity than those with a negative PCR and a negative LST. The most important finding was that, during follow-up, eight (57%) of the PCR-positives, but none of the 21 negatives, developed post-kala-azar dermal leishmaniasis (PKDL) In conclusion, PCR-based testing of lymph-node aspirates after treatment may be used as a prognostic marker for the future development of PKDL and may be useful in the follow-up of patients.

Agglutination Tests↗
PubMed1997

Evaluation of PCR for diagnosis of visceral leishmaniasis.

An evaluation of Leishmania PCR was performed with bone marrow, lymph node, and blood samples from 492 patients, 60 positive controls, and 90 negative controls. Results were compared with microscopy results for Giemsa-stained smears. PCR and microscopy of lymph node and bone marrow aspirates from patients with microscopically confirmed visceral leishmaniasis (VL) were equally sensitive. However, in patients clinically suspected of having VL and in whom parasites could not be demonstrated by microscopy, PCR was positive for 12 of 23 (52.2%) lymph node aspirates and 8 of 12 (66.7%) bone marrow aspirates, thus confirming the clinical diagnosis of VL. With PCR on filter paper, Leishmania DNA was detected in the blood of 33 of 47 (70%) patients with confirmed VL and in 2 of 11 (19%) patients suspected of having VL. Positive PCR results were more frequently found for blood samples on filter paper than for samples stored in EDTA. In conclusion, PCR is a more sensitive method than microscopy for the detection of Leishmania in lymph node and bone marrow aspirates, being especially useful for the confirmation of cases of suspected VL. Blood from a finger prick may be used for the initial PCR screening of people suspected of having VL. If the PCR of blood is negative, one should perform PCR with lymph node and/or bone marrow material, because PCR with these materials is more often positive.

Blood↗
PubMed1996

Identification and determination of the relationships of species and strains within the genus Leishmania using single primers in the polymerase chain reaction.

DNA polymorphisms were assessed in different species and strains within the genus Leishmania by amplifying genomic DNA with single non-specific primers. This polymerase chain reaction (PCR) method employed non-random primers which anneal to mini- and microsatellite DNA sequences like the M13 core sequence and the simple repeat sequences (GTG)5 and (GACA)4, and the T3B primer derived from an intergenic spacer for tRNA genes. Distinctive and reproducible sets of amplified DNA fragments were obtained for all Leishmania isolates tested. The number and size of amplification products were found to be characteristic for a given taxon. Highly similar PCR profiles were observed when genomic DNA of representatives of the L. donovani, L. mexicana or L. braziliensis complexes was amplified. By comparing PCR patterns of unidentified Leishmania isolates with those obtained from reference strains it was possible to identify these isolates at the species level. The information of the amplification patterns was used for the construction of phylogenetic trees to measure the genetic relatedness within the genus Leishmania.

Animals↗
PubMed1996

Polymerase chain reaction for detecting Onchocerca volvulus in pools of blackflies.

The detection of Onchocerca volvulus infected simuliids or blackflies is routinely done by dissection and microscopic examination of individual flies, but this method is tedious and time consuming. Here we describe a method of detecting single O. volvulus infected blackflies in pools of uninfected blackflies. Using a PCR with Onchocerca specific primers it is possible to reproducibly detect one heavily infected blackfly in a pool of 80 flies, or to detect one blackfly inoculated with one microfilaria in a pool of 20 flies. With the method described large numbers of blackflies can be rapidly screened for the presence of O. volvulus infected flies.

Animals↗
PubMed1995

Epidemiology, diagnosis and control of leishmaniasis in the Mediterranean region.

The leishmaniases are a widespread and medically important group of parasitic diseases, some of which pose a serious health threat in communities throughout the Mediterranean basin. In 1993, a joint, collaborative study of the Mediterranean leishmaniases was initiated by scientists from Israel, Turkey, Portugal and the Netherlands. The aim of this project was the development of a multi-component approach to the successful control of all forms of leishmaniasis, with special emphasis on the more severe, visceral leishmaniasis (VL). The need for highly sensitive and accurate new tools to facilitate diagnosis and epidemiological surveys of endemic areas and for studies on the immunology of VL in laboratory models (dogs and mice) was soon recognized. It is anticipated that the development of these tools and the associated technology will provide a better understanding of the disease and improve its control.

Animals↗
PubMed1995

Leish-KIT, a stable direct agglutination test based on freeze-dried antigen for serodiagnosis of visceral leishmaniasis.

In order to increase the application potential of the direct agglutination test (DAT) for the detection of anti-Leishmania antibodies in human serum samples, we developed an antigen based on stained and freeze-dried Leishmania donovani promastigotes. We describe here the evaluation of the performance of the DAT based on this freeze-dried antigen. It was shown that the freeze-dried antigen remains fully active, even after storage at 56 degrees C for 18 months. With a cutoff value of 1:1,600, the sensitivity of the DAT was shown to be 92% and the specificity of the test was 99.7%, which were comparable with the results found for the DAT based on liquid antigen. The major advantages of the freeze-dried antigen are that the production of a large batch of this antigen allows reproducible results in the DAT over a long period of time and that the freeze-dried antigen can be stored at ambient temperature, which, as was shown, makes the test a valuable diagnostic tool for use in the field.

Agglutination Tests↗
PubMed1992

Plasmid maintenance in Bacillus stearothermophilus is strain-dependent.

We studied the segregational stability of plasmids based on pTB913, a 4.5-kb rolling-circle plasmid derived from the thermophilic Bacillus plasmid pTB19. In Bacillus stearothermophilus the stability of pTB913 derivatives appeared to be strain-dependent. In strain CU21 large amounts of single-stranded pTB913 DNA were found and the plasmid was highly unstable at 57 degrees C. In strain NUB3621, however, very low amounts of single-stranded plasmid DNA were formed and pTB913-based replicons were only slightly unstable at 57 degrees C. The NUB3621/pTB913 host-vector system seems appropriate for molecular cloning. A RepA-based replicon, also derived from pTB19 but replicating by a theta mechanism, was highly unstable in B. stearothermophilus NUB3621.

DNA Replication↗
PubMed1992

The integrated state of the rolling-circle plasmid pTB913 in the composite Bacillus plasmid pTB19.

pTB19, a 27 kb plasmid originating from a thermophilic Bacillus species, contains integrated copies of two rolling-circle type plasmids on a 10.6 kb DNA fragment. In the present study we analysed the part of pTB19 that contains the rolling-circle plasmid pTB913 and the region in between the two rolling-circle plasmids. We show that, in the integrated state, pTB913 was flanked by a 55 bp direct repeat that duplicated part of the replication initiation gene repB. Since repB was interrupted, the integrated pTB913 could not initiate rolling-circle replication. Autonomously replicating pTB913 was produced from pTB19, probably through recombination between the 55 bp direct repeats; this was a rare event. Since the second integrated rolling-circle type plasmid also contained a non-functional replication initiation gene, replication of pTB19 must be controlled by the RepA determinant. Theta-type replication, controlled by RepA is likely to account for the high stability of pTB19. In between the two integrated rolling-circle plasmids was present an open reading frame (447 codons) which could encode a protein of unknown function.

Amino Acid Sequence↗
PubMed1992

Plasmid pTB913 derivatives are segregationally stable in Bacillus subtilis at elevated temperatures.

We studied the effects of temperature on the segregational stability of derivatives of the rolling-circle-type plasmid pTB913 in Bacillus subtilis. This 4.5-kb plasmid is a deletion derivative of pTB19, which was originally isolated from a thermophilic Bacillus. pTB913 derivatives carrying large inserts or lacking the minus origin for complementary strand synthesis were segregationally unstable at 37 degrees C. In contrast, at 47 degrees C all pTB913 derivatives tested were stably maintained in B. subtilis. The increased stability at 47 degrees C was attributed, at least partly, to increased copy numbers at this temperature. Although considerable amounts of single-stranded and high-molecular-weight plasmid DNA were formed at 47 degrees C, these products did not reduce plasmid stability at this temperature. The increased stability and increased copy number of pTB913 at elevated temperatures extend the use of this plasmid as a cloning vector in B. subtilis and other bacilli.

Bacillus subtilis↗
PubMed1991

The large Bacillus plasmid pTB19 contains two integrated rolling-circle plasmids carrying mobilization functions.

Plasmid pTB19 is a 27-kb plasmid originating from a thermophilic Bacillus species. It was shown previously that pTB19 contains an integrated copy of the rolling-circle type plasmid pTB913. Here we describe the analysis of a 4324-bp region of pTB19 conferring resistance to tetracycline. The nucleotide sequence of this region revealed all the characteristics of a second plasmid replicating via the rolling-circle mechanism. This sequence contained (i) the tetracycline resistance marker of pTB19, which is highly similar to other tetL-genes of gram-positive bacteria; (ii) a hybrid mob gene, which bears relatedness to both the mob-genes of pUB110 and pTB913; (iii) a palU type minus origin identical to those of pUB110 and pTB913; and (iv) a plus origin of replication similar to that of pTB913. A repB-type replication initiation gene sequence identical to that of pTB913 was present, which lacked the middle part (492 bp), thus preventing autonomous replication of this region. The hybrid mob gene was functional in conjugative mobilization of plasmids between strains of Bacillus subtilis.

Amino Acid Sequence↗
PubMed1990

Conjugal mobilization of streptococcal plasmid pMV158 between strains of Lactococcus lactis subsp. lactis.

pMV158, a non-self-transmissible plasmid encoding tetracycline resistance, was conjugally transferred from Enterococcus faecalis JH203 to Lactococcus lactis subsp. lactis IL1403. This transfer appeared to be dependent on the cotransfer of the conjugative plasmids pAM beta 1 or pIP501. Intraspecies conjugal transfer of pMV158 also occurred in strain IL1403. In contrast to the transfer from E. faecalis, transfer in IL1403 did not require the presence of a conjugative plasmid in the donor strain but, rather, appeared to be dependent on putative chromosomal functions in strain IL1403. The transfer of pMV158 from strain IL1403 required the presence of an active pMV158-encoded protein, which showed homology to the Pre (plasmid recombination enzyme) proteins encoded by several small plasmids extracted from Staphylococcus aureus, such as pT181.

Conjugation, Genetic↗
PubMed1989

Similarity of minus origins of replication and flanking open reading frames of plasmids pUB110, pTB913 and pMV158.

Plasmids pMV158 and pTB913, originating from Streptococcus agalactiae and a thermophilic Bacillus respectively, were sequenced to completion. Both contained a BA3-type minus origin of replication and an RSA-site, believed to constitute a site-specific recombination site. These two regions were more than 99% homologous to the corresponding regions of the Staphylococcus aureus plasmid pUB110. Deleting the BA3-type minus origin resulted in the accumulation of a considerable amount of single-stranded DNA, both in L. lactis subsp. lactis and B. subtilis, indicating that this minus origin was functional in both bacterial species. Like pUB110, both plasmids contained an open reading frame encoding a putative plasmid recombination enzyme (Pre protein), which was located downstream of the RSA-site. On the basis of sequence comparisons between pUB110, pMV158, pTB913, pT181, pE194, pNE131 and pT48 two distinct families of RSA-sites and Pre proteins could be distinguished.

Amino Acid Sequence↗
PubMedDate not supplied

Genetic heterogeneity in the species Leishmania tropica revealed by different PCR-based methods.

A PCR fingerprinting approach, using single non-specific primers, as well as restriction and single-strand conformation polymorphism (SSCP) analyses of the amplified ribosomal internal transcribed spacer, were used to investigate genetic variability in the species Leishmania tropica. Twenty-nine strains of the 'L. tropica complex' from different Old World geographical areas were studied including 4 from Namibia, and 1 strain of L. killicki. All techniques revealed a high degree of genetic heterogeneity among the strains of L. tropica. The PCR fingerprinting displayed the highest discriminatory power, but can be applied only to cultured parasites. The internal transcribed spacer (ITS) region can be amplified directly from infected clinical samples and analysed subsequently. No strict correlation was discerned between the genetic variants and either the geographical origin of the strains or the clinical manifestations associated with human disease, except for the Namibian strains. Also, genetic variation did not correlate well with characterization by enzyme variant electrophoretic analysis. The strain of L. killicki always clustered together with the strains of L. tropica, suggesting it, probably, should not be considered a separate species of Leishmania. However, the 4 Namibian strains formed a distinct, statistically well-supported group closely related to but different from the other strains of L. tropica.

Animals↗
PubMedDate not supplied

The direct agglutination test for diagnosis of visceral leishmaniasis under field conditions in Sudan: comparison of aqueous and freeze-dried antigens.

The performance of the direct agglutination test (DAT) was evaluated under field conditions in an endemic area of visceral leishmaniasis in eastern Sudan, using aqueous (Aq) antigen which has to be kept refrigerated and a newly developed freeze-dried (FD) antigen which is stable at ambient temperature. Both antigens compared well, with 92-98% of readings being identical or only with one dilution difference in titre. FD antigen gave titres that were identical with Aq antigen in 73% of samples, higher in 19%, and lower in 8%. Owing to high ambient temperatures and low humidity, microtitre plate wells dried out during the standard procedures for elution and incubation. However, shortening the elution time from 12 to 4 h proved possible for both antigens; incubation could be reduced from 24 to 10 h for Aq antigen, after which the plates could still be read. Incubation with FD antigen required 18 h and the plates needed to be kept cool because of evaporation. Despite the longer procedure with the FD antigen, the DAT can be completed in 24 h and the use of this stable antigen, that does not require refrigeration, is a major improvement in performing the DAT under unfavourable field conditions.

Agglutination Tests↗
PubMedDate not supplied

Biochemical and molecular characterization of Leishmania parasites isolated from an endemic focus in eastern Sudan.

Twelve Leishmania isolates from visceral leishmaniasis patients in eastern Sudan were characterized using isoenzyme analysis, Southern blotting and polymerase chain reaction (PCR) 'fingerprinting'. Isoenzyme analysis revealed the presence of 3 zymodemes: MON-18, MON-30 and MON-82, corresponding to Leishmania donovani sensu stricto, L. infantum and L. archibaldi (still of uncertain taxonomic status), respectively. Southern blotting and PCR 'fingerprinting' revealed identical patterns for all 3 zymodemes, which were indistinguishable from those of L. donovani s.s.

Animals↗