Electronic properties of site-controlled (111)-oriented zinc-blende InGaAs/GaAs quantum dots calculated using a symmetry-adapted k·p Hamiltonian.

In this work, we present and evaluate a (111)-rotated eight-band k ⋅p Hamiltonian for the zinc-blende crystal lattice to investigate the electronic properties of site-controlled InGaAs/GaAs quantum dots grown along the [111] direction. We derive the rotated Hamiltonian including strain and piezoelectric potentials. In combination with our previously formulated (111)-oriented continuum elasticity model, we employ this approach to investigate the electronic properties of a realistic site-controlled (111)-grown InGaAs quantum dot. We combine these studies with an evaluation of single-band effective mass and eight-band k ⋅p models, to investigate the capabilities of these models for the description of electronic properties of (111)-grown zinc-blende quantum dots. Moreover, the influence of second-order piezoelectric contributions on the polarization potential in such systems is studied. The description of the electronic structure of nanostructures grown on (111)-oriented surfaces can now be achieved with significantly reduced computational costs in comparison to calculations performed using the conventional (001)-oriented models.

Establishment of a new bovine leukosis virus producing cell line.

Due to the prevalence of different bovine leukosis virus (BLV) species in the cattle population in Europe, problems may arise in the serological diagnosis of BLV infections. In addition, earlier investigations demonstrated that contamination of the BLV antigen-producing cell culture systems by bovine viral diarrhea virus (BVDV) may give rise to misinterpretation of serological test results after BVDV vaccination of cattle. By co-cultivation of peripheral leukocytes of a BLV-infected cow with a permanent sheep kidney cell line, a new BLV-producing cell line named PO714 was established. This line carries a BLV provirus of the Belgian species and has been tested to be free of a variety of possibly contaminating viruses and mycoplasms. Investigations of a panel of well-characterised sera by agar gel immunodiffusion (AGID) and capture ELISA (cELISA) tests using antigen prepared from this new cell line in comparison with antigen of the well-known cell line FLK/BLV yielded comparable results. False positive results caused by BVDV cross-reactions could be eliminated when tests were carried out with antigen derived from the new cell line.

Differentiation of type A, Asia1 and O foot-and-mouth disease virus variants, amplified by the same system, by sequencing of the capsid protein genes.

A reverse transcription-dependent polymerase chain reaction (RT-PCR) is described that amplifies the genes encoding the capsid proteins VP1-3 of at least three evolutionary lineages each of the foot-and-mouth disease (FMD) virus types A, Asia1 and O. Most of these lineages are circulating at present in Asia and Africa. The method is not only suitable to confirm suspected outbreaks of FMD, but also describes the modulation of major and minor antigenic sites in the course of an epizootic by nucleotide sequence determination of the obtained RT-PCR products. Such knowledge helps to choose suitable vaccines for disease control.

An outbreak of enzootic bovine leukosis in upper Egypt: clinical, laboratory and molecular-epidemiological studies.

In 1989, 220 Holstein Friesian cattle (212 heifers and eight bulls) were imported from Minnesota, USA, to form a closed dairy herd in Arab El-Aoumar, Assiut, Upper Egypt. In November 1996, some abnormal signs such as loss of weight, decreased milk yield, external lymphadenopathy and decreased appetite were observed on this farm. Serological screening by enzyme-linked immunosorbent assay revealed a seroprevalence of antibodies directed against bovine leukaemia virus (BLV) of 37.7% in cattle under 2 years old and of 72.8% in animals more than 2 years old. Diagnosis was confirmed by the detection of BLV proviral DNA using polymerase chain reaction with primers amplifying a fragment of the env gene. Out of 21 tested leucocyte fractions from individual animals, 15 were positive showing a BLV-specific amplicon of 444 base pairs. Analysis of the amplicons for restriction fragment length polymorphisms and DNA sequencing results allowed the isolates to be typed. Since this was the first recorded case of enzootic bovine leukosis in Upper Egypt, strict quarantine measures were adopted and all serologically positive animals in the herd were culled.

Epidemiological implications of the molecular characterization of foot-and-mouth disease virus isolated between 1996 and 2000 in Bangladesh.

Foot-and-mouth disease virus was collected during two years throughout Bangladesh. Viral RNA from 40 samples was subjected to reverse transcription-dependent polymerase chain reactions that amplify parts of the capsid protein encoding genome region, and the products obtained were sequenced. This showed that all virus isolates up to January 1999 belonged to a genotype of serotype O, observed here already in 1987, 1996 and 1997, and elsewhere since 1990. In February 2001, this virus variant was introduced into Great Britain and then transmitted to other European countries. The capsid protein sequences of an isolate of 2001 from the Netherlands is provided. Later isolates from Bangladesh, however, belonged to a genotype of serotype A that had been transmitted to Albania in 1996. No virus of type Asia1 was found, although it circulated in Bangladesh in 1996. Instead, this genotype of Asia1 virus was observed in Iran late in 1999, and transmitted from Turkey to Greece in July 2000. The results indicate continued intercontinental transmission of foot-and-mouth disease viruses that circulate in central Asia.

Identification of different BLV provirus isolates by PCR, RFLPA and DNA sequencing.

A first attempt for the investigation of molecular epidemiology of BLV was carried out. PCR amplicons of a part of the env gene of BLV isolated from 309 cattle of different geographical origin were compared with known BLV env sequences. Using RFLPA most of the PCR products can be assigned to the Australian, the Japanese or the Belgian subgroup. A phylogenetic tree resulting from the comparison of the sequences of these env fragments demonstrates the relations and differences between and within the subgroups.

Molecular epidemiology of serotype O foot-and-mouth disease virus with emphasis on West and South Africa.

Genetic relationships of serotype O foot-and-mouth disease (FMD) viruses recovered from outbreaks of the disease in the West African countries of Niger, Burkina Faso and, Ghana (1988-1993) and those from South Africa (2000) were determined by partial VP1 gene characterization. A 581-bp fragment, corresponding to the C-terminus half of the ID (VP1 gene) region was amplified and sequenced. An homologous region of 495 nucleotides was ultimately used to determine genetic relationships of serotype O viruses from the Middle East, Europe, South America, North Africa, East Africa, southern Africa and Asia. Seven distinct type O genotypes were identified by phylogenetic reconstruction, consisting of viruses from the following geographical regions: Genotype A: Asia, the Middle East, and South Africa, Genotype B: East Africa, Genotype C: West and North Africa, Genotype D: Taiwan and Russia, Genotype E: Angola and Venezuela, Genotype F: Western Europe, and Genotype G: Europe and South America. The genotypes constitute three different evolutionary lineages (I-III), which correspond to three discrete continental regions, some of which display inter-continental distributions due to introductions. Results further indicate that the outbreaks in Burkina Faso (1992) and Ghana (1993) are part of the same epizootic and that the strain involved in a recent outbreak of the disease in South Africa is most closely related (97% sequence identity) to a 1997 Bangladesh strain.

Type-independent detection of foot-and-mouth disease virus by monoclonal antibodies that bind to amino-terminal residues of capsid protein VP2.

The characterization of monoclonal antibodies raised against the foot-and-mouth disease virus isolates A22 Iraq/1964, Asia1 Shamir-Israel/1989, and SAT1 Zimbabwe/1989 with regard to neutralizing activity and sensitivity of their epitopes for treatment with trypsin, resulted in the identification of one non-neutralizing antibody in each panel that binds to a trypsin-sensitive epitope. Furthermore, each of these antibodies recognized 27 isolates of different provenance, representative of six serotypes. These antibodies are recommended for type-independent antigen detection by ELISA. The epitopes for these antibodies reside at the intertypically conserved N-terminus of capsid protein VP2. The two are specified by the lysines at positions two and three, but differ from each other as indicated by the variable heavy chain sequences of their antibodies.

Nucleotide sequence, genomic organization and cell-cycle-dependent expression of a Chlamydomonas 14-3-3 gene.

Members of the 14-3-3 protein family have been identified as regulatory elements in intracellular signalling pathways and cell cycle control. Previously we reported the nucleotide sequence of a 14-3-3 cDNA cloned from the unicellular green alga Chlamydomonas reinhardtii. In this communication, we describe the nucleotide sequence, the genomic organization and the cell-cycle-dependent expression of the corresponding gene. The coding sequence of this gene was found to be interrupted by four introns of 124, 116, 81, and 659 bp, respectively. Introns 2-4 were found in conserved positions as compared to the Arabidopsis 14-3-3 genes. A counterpart to intron 1 absent in the Arabidopsis 14-3-3 genes was found in the human 14-3-3 epsilon gene.

A nucleotide deletion causing a translational stop in the protease reading frame of bovine leukaemia virus (BLV) results in modified protein expression and loss of infectivity.

Bovine leukaemia virus (BLV) is an oncogenic retrovirus that causes B-cell lymphocytosis and in the terminal stage of the disease lymphosarcoma. The comparison of the previously published BLV provirus sequence from Belgium, Australia and Japan showed that the protease gene (prt) of the Australian and the Japanese isolate contain a nucleotide deletion when compared to the Belgian isolate. Because all these proviruses were isolated from tumour tissue, the prt gene of functionally active and infectious proviruses from peripheral blood leucocytes (PBLs) of BLV-infected cattle and from BLV-infected fetal lamb kidney cells were sequenced. The only variations between these sequences and the Belgian isolate consist of nucleotide substitutions. The delection of one nucleotide of the prt gene of the Japanese and the Australian BLV tumour isolate caused a changed reading frame and a premature translational stop. It was shown that the Japanese provirus is non-infectious in transfected cell culture and in injected sheep. To analyse the impact of the prt mutation on viral protein expression and infectivity, the prt region of the Japanese provirus was exchanged with the prt region from the Belgian provirus. The resulting pBLVprtbelg was infectious in transfected cells and enabled the expression of gag and gag-precursor proteins. One sheep was injected with this mutated provirus and became positive in BLV-PCR, but no seroconversion was developed. The prt mutation of the Japanese tumour isolates was shown to be responsible for the loss of infectivity and changed viral expression. These results and the occurrence of this mutation in only two isolates from lymphosarcoma indicate a possible relation between the prt mutation and the induction of cell transformation.

Antigenic variation among foot-and-mouth disease virus type A field isolates of 1997-1999 from Iran.

The sequences of the antigenically relevant capsid proteins VP1-3 of 10 isolates obtained during an epizootic of serotype A foot-and-mouth disease virus in Iran, and collected within two and a half years, were found to be highly similar. However, each isolate differed by at least one amino acid from all others. This prompted us to analyze the immunological reactivity of the isolates. To this end, monoclonal antibodies (mAbs) against one isolate were generated and characterized with regard to neutralizing activity and reactivity with trypsinized virus. These mAbs as well as others raised against A22 virus were used for antigen profiling. This distinguished four antigenic conditions among the isolates and 16 reactivities among the mAbs. These findings, together with the observed sequence differences indicated the location of several epitopes. Many mAbs recognized the minor antigenic sites on VP2 and 3 and some the major site, the GH-loop of VP1. One epitope was composed of residues of the capsid proteins VP1 and 2.

Genetic and antigenic variance of foot-and-mouth disease virus type Asia1.

The capsid protein encoding genes of five recent type Asia1 foot-and-mouth disease virus isolates, representative of three genotypes, were sequenced. The deduced amino acid sequences were aligned to each other and to two published sequences. The sequence differences suggested different antigenic properties of the isolates. One isolate was used to generate monoclonal antibodies (mAbs) which were analyzed for neutralizing activity and reactivity with trypsinized virus. Trypsin removes the major antigenic sites located at VP1. The five virus isolates formed three reaction patterns with the mAbs, irrespective of their genotype. Combination of all data allowed to suggest the location of the epitope of each antibody: the VP1 G-H and the VP2 B-C loop, the VP3 B-B knob, and the N-terminus of VP2, respectively, were involved.

Genetical and immunological analysis of recent Asian type A and O foot-and-mouth disease virus isolates.

This report extends the knowledge on the epizootical situation of foot-and-mouth disease in Asia. RNA from six samples of type A and five of type O virus, isolated between 1987 and 1997 in Bangladesh, Iran, Malaysia and Turkey, was subjected to reverse transcription-dependent polymerase chain reactions that amplify large parts of the capsid protein VP1 encoding genome region. The amplification products were sequenced, and the sequences aligned to each other and to published sequences. This showed the type O isolates of 1987-1997 from Bangladesh to be of same genotype and closely related to isolates of 1988 and later from Saudi Arabia, 1990 from India, 1996 from Greece and Bulgaria, and 1997 from Iran. Among the analyzed type A isolates, those of 1992 and 1996 from Turkey were of same genotype and related to previously described isolates of 1987 from Iran and of 1992 from Saudi Arabia. The isolate of 1997 from Malaysia was found to be related to isolates from Thailand of 1993 and 1996. The isolates of 1987 from Bangladesh and 1997 from Iran, however, represent different so far not described genotypes. Monoclonal antibodies, raised against the vaccine production strains A22 Iraq, Asial Shamir, O1 Kaufbeuren and O1 Manisa, and the recent type A field isolates Saudi Arabia/92 and Albania/96, were used in an ELISA to compare the reaction patterns of many of the field isolates. The monoclonal antibodies were further characterized for virus-neutralizing activity and binding to trypsinized homologous virus. The failure of neutralizing antibodies in binding to trypsinized homologous as well as to heterologous virus suggested the epitopes to reside at the major antigenic component of the virus, which is the capsid protein VP1. Two non-neutralizing antibodies that bind to trypsin-sensitive epitopes cross-reacted, however, with heterologous virus. This indicates the existence of a trypsin-sensitive antigenic site outside of VP1. In summary, the results obtained by ELISA confirm the observed sequence differences, but indicate further sequence differences at minor antigenic sites that do not reside on VP1.

[Serologic and virologic investigations on the presence of BLV infection in a dairy herd in Syria]. / Serologische und virologische Untersuchungen über das Vorkommen der BLV-Infektion in einer Milchviehherde in Syrien.

237 cattle of a dairy herd in Syria were tested for anti-BLV antibody by the ELISA. 194 animals were additionally examined by the agar gel immunodiffusions test (AGID) on BLV antibodies and 100 by polymerase chain reaction (PCR) for BLV provirus. BLV specific antibodies were determined by means of AGID and ELISA at 62.9% and 69.2% of the examined animals, respectively. Using the PCR method the BLV provirus was detected in 89% of the investigated cattle. Only one ELISA seropositive animal was negative for BLV provirus. The results show the high BLV contamination of this herd and lead to the presumption of wide spread enzootic bovine leukosis in Syria. In the case of the diagnosis of BLV-infection, the PCR-technique compared to the serological tests proved to be much more sensitive. By the detection of BLV antibody, the ELISA showed a higher sensitivity than the AGID and in this way, is advisable as a method of choice for screening investigations. Restriction enzyme and sequence analysis of PCR-amplificates demonstrate that different BLV provirus variants (A, B and C) in the examined herd occur, where the variant C which a high similarity to an Australian BLV provirus isolates showed, occurred most frequently at 92.5%.

Quantitation of foot-and-mouth disease virus genomes in bovine tissue by competitive RT-PCR.

The sensitivity of a reverse transcription-dependent polymerase chain reaction (RT-PCR) for detecting foot-and-mouth disease virus (FMDV) genomes was quantified by use of RNA transcribed in vitro from FMDV-specific cDNA. Previously, the cDNA had been elongated by 228 base pairs. The minimum number of template molecules required to obtain the specific RT-PCR product was determined to be 10(4). This was achieved by use of 1 microg of primer for cDNA synthesis and by undertaking of at least 30 cycles of PCR. Knowing the sensitivity of the system prompted the examination of clinical samples for content of FMDV genomes. The samples were tongue and foot epithelia as well as nasal discharge, removed 11-14 days after infection from 14 cattle. They all contained FMDV genomes but not in each clinical specimen. The size difference of the products amplified from transcript and viral genome enabled the estimate by competitive RT-PCR of the number of viral genomes contained in some samples. The RNA extracted from approximately 10(7) tissue cells each was found to contain between less than 10(6) and up to 10(8) FMDV genomes, irrespective of the sample type.

VP1-coding sequences of recent isolates of foot-and-mouth disease virus types A, O and Asia1.

A large part of the capsid protein VP1-coding sequence of foot-and-mouth disease virus, isolated between 1993 and 1996 in Europe, was amplified by the reverse transcription-dependent polymerase chain reaction (RT-PCR). The same was done with some non-European virus isolates, especially those against which vaccines were currently produced. The products were sequenced, and the sequences aligned. The alignment comprises sequences of the types A, O and Asia 1. Although the provenance of virus introduced to Europe remains unknown, genetic relation to some other isolates was indicated. Several genotypes of the virus were found to circulate in the field since years.

[Polymerase chain reaction (PCR) for detection of BLV provirus-- a practical complement for BLV diagnosis?]. / Polymerase Kettenreaktion (PCR) zum Nachweis von BLV-Provirus--praktikable Ergänzung der BLV-Diagnostik?

A typical infection with bovine leukemia virus (BLV) induces a permanent antibody (Ab) response with high titers against BLV-antigens. In the last few years atypical courses of infection with low or transient BLV-Ab-titers or even lack of any detectable BLV-Ab-titers in animals with BLV-provirus integration have been described. This makes it difficult to eliminate BLV infection from herds using serological assays only. Whether or not polymerase chain reaction (PCR) is a useful tool to complement serological Ab-assays in BLV-eradication in herds was clarified in three ways: (i) different DNA-quick-preparations of blood were examined in nested PCR, (ii) cows of a BLV infected herd that was involved in a national eradication program were investigated for 6 months und (iii) BLV-provirus-variants occurring in this herd were differentiated. The results show, that even by using PCR it was not possible to detect all infected animals all the time and that eradication of BLV from this herd was not completed in this short time. The PCR is useful for the investigation of herds and more sensitive than ELISA. PCR using LTR-primers (34 positive cattle) was more sensitive than PCR with env-primers (30 positive cattle). Using PCR 34 BLV infected cattle were detected of which only 21 reacted in ELISA. Restriction enzyme analysis or sequence analysis of PCR-amplificates allowed the detection of virus variants and conclusions about the way of infection. PCR should be used for BLV-eradication in cattle herds with low BLV-incidence, for the investigation of new outbreaks or tumor cases in long term BLV free herds and for investigation of breeding cattle.

Provirus variants of the bovine leukemia virus and their relation to the serological status of naturally infected cattle.

Infection of cattle with the bovine leukemia virus (BLV) results in a strong permanent antibody response to the BLV antigens some weeks after infection. However, cattle may carry provirus and not have detectable antibody titers. To prove the occurrence of different BLV provirus variants in German cattle and to study the influence of special BLV variants on the immunoreaction, a 444-bp fragment of the env gene of 35 naturally BLV infected animals was analyzed. Seven different groups of BLV provirus variants were found on the basis of restriction fragment length polymorphism. Three BLV provirus variant groups and five additionally sequenced BLV isolates showed a high similarity to BLV provirus isolates from other geographical areas. The variation in nucleotide sequence of the five BLV isolates compared with nine previously sequenced BLV isolates ranged up to 5. 3%. While BLV provirus variant groups A, C, D, E, F, and G were clearly related to agar-gel immunodiffusion test (AGID)- and enzyme-linked immunosorbent assay (ELISA)-positive animals, BLV provirus variant group B was solely found in permanent AGID- and ELISA-negative or in transient ELISA-positive animals. Altogether, these results indicate that special BLV provirus variants may be responsible for atypical forms of BLV infection in cattle.