Miklós Kálmán

Microbial population in a hydrogen-dependent denitrification reactor.

The bacterial population in an H2-dependent denitrification system was studied. The laboratory set-up was designed for the treatment of potable water and consisted of an electrochemical cell, where the water to be treated was enriched with H2 prior to entering a bioreactor. Bioreactors (columns packed with granulated active carbon) were inoculated with denitrifying bacterial strains isolated from a previous reactor, then sampled immediately after inoculation, or after 1 or 3 months of continuous operation. Total number of the bacteria and numbers of each different strain were determined at various levels of the bioreactor. The strains present in the inoculum were identified as Ochrobactrum anthropi, Pseudomonas stutzeri, Paracoccus panthotrophus and Paracoccus denitrificans. Numbers of the latter declined markedly with time with the other three strains being responsible for nitrate removal. A correlation was found between the relative abundance of each strain and its specific denitrification activity.

The metD d-Methionine Transporter Locus of Escherichia coli Is an ABC Transporter Gene Cluster

The metD D-methionine transporter locus of Escherichia coli was identified as the abc-yaeE-yaeC cluster (now renamed metNIQ genes). The abc open reading frame is preceded by tandem MET boxes bracketed by the -10 and -35 boxes of a promoter. The expression driven by this promoter is controlled by the MetJ repressor and the level of methionine.

Milkborne campylobacter infection in Hungary.

In April 1998, an annual 2-day animal farm sale was held in Hódmezóvásárhely, where 500 to 600 visitors consumed unpasteurized milk. The first signs of disease began 2 days after the end of the sale. Fifty-two people from a wide age range fell ill, primarily with inflammatory enteritis. These cases included 34 with Campylobacter positivity: 30 with Campylobacter jejuni and 4 with Campylobacter coli. Environmental samples (raw milk, udder swabs, and rectal swabs from 12 cows in the suspected herd) were tested 2 weeks after the first signs of the disease, and two rectal swabs were found to be positive for C. jejuni. Initially, the epidemic seemed to be sporadic and, accordingly, only 26 human and 2 animal Campylobacter isolates were reserved for randomly amplified polymorphic DNA analysis. This comparative analysis verified that fecally contaminated milk was the source of the outbreak. The DNA-banding patterns of 20 C. jejuni isolates (19 human and 1 animal) were identical. The antibiotic susceptibilities of the Campylobacter isolates were determined, and only six C. jejuni (human) isolates, one C. coli (human) isolate, and one C. jejuni (animal) isolate were resistant to tetracycline, both by disk diffusion and by E test (antimicrobial gradient strip for the quantitative determination of susceptibility or resistance of microorganisms). No plasmid was detected in these tetracycline-resistant isolates. The endotoxin production of Campylobacter isolates was examined via the cytopathogenic effect on the Vero cell line. This effect exhibited various degrees of positivity in 19 cases. Only two human C. jejuni isolates displayed + + + + positivity. Both isolates were from patients who had required antibiotic therapy and hospital care.

Directional cloning of native PCR products with preformed sticky ends (autosticky PCR).

Abstract A novel method for the directional cloning of native PCR products was developed. Abasic sites in DNA templates make DNA polymerases stall at the site during synthesis of the complementary strand. Since the 5′ ends of PCR product strands contain built-in amplification primers, abasic sites within the primers result in the formation of 5′ single-stranded overhangs at the ends of the PCR product, enabling its direct ligation to a suitably cleaved cloning vector without any further modification. This “autosticky PCR” (AS-PCR) overcomes the problems caused by end sensitivity of restriction enzymes, or internal restriction sites within the amplified sequences, and enables the generation of essentially any desired 5′ overhang.

Characterization of the Escherichia coli K12 gltS glutamate permease gene

SummaryThe gltS gene is known to encode a sodium-dependent, glutamate-specific permease. We have localized the Escherichia coli K12 gltS gene with respect to the spoT gene, sequenced it, and recombined a null insertion-deletion allele into the chromosome without loss of viability. The gltS null allele gives a Glt− phenotype, i.e. it abolishes the ability of a gltCc host to grow on glutamate as sole carbon and nitrogen source and also confers α-methylglutamate resistance. A multicopy plasmid expressing the gltS gene can reverse the Glt− phenotype of gltS− or wild-type strains while other plasmids show host-dependent complementation patterns. Induction of gltS gene overexpression under control of isopropylβ-d-thiogalactoside (IPTG)-inducible promoters severely inhibits growth. The G1tS protein is deduced to be a 42425 dalton hydrophobic protein with 2 sets of 5 possible integral protein domains, each flanking a central hydrophilic, flexible region.

Polymerase chain reaction (PCR) amplification with a single specific primer.

A method is described for amplification of DNA fragments flanking a single known sequence that is sufficiently long to enable synthesis of a functional primer in polymerase chain reactions.

Recent advances in biohydrogen research

Abstract. A fundamental and principal difficulty of the future energy supply is that the formation of fossil fuels is much slower than the rate of their exploitation. Therefore the reserves which can be recovered in an energetically feasible manner are shrinking parallel with an increasing world-wide energy demand. Among the alternative energy carriers, hydrogen is preferred because it is easy to transport and store and it burns to environmentally friendly water vapour when utilized. Hydrogen can be produced in biological systems, however, our understanding of the molecular details is just emerging.

A novel polyacrylamide-type support prepared by p-benzoquinone activation.

A new and simple method for the activation of polyacrylamide gels usingp-benzoquinone is described. The optimal conditions of activation have been elaborated. The activated support could be successfully applied to the immobilization of ligands having nucleophilic groups active over a broad pH range.

Functional EF-Tu with large C-terminal extensions in an E. coli strain with a precise deletion of both chromosomal tuf genes

An Escherichia coli strain was constructed in which both chromosomal genes encoding elongation factor (EF)‐Tu (tufA and tufB) have been inactivated with precise coding sequence replacements. A tufA gene in an expression vector is supplied as the sole EF‐Tu source. By using plasmid replacement, based on plasmid incompatibility, mutant EF‐Tu variants with a large C′‐terminal extension up to 270 amino acids were studied and proved to be functional in a strain lacking the chromosomal tufA and tufB genes.

Effect of phytohaemagglutinin on CD45 in T cells

In this study the effect of PHA activation on the phosphatase activity of CD45 has been investigated in human leukemic T-cell lines. It has been found that in vivo activation of the cells with PHA resulted in 2-4-fold increase in enzyme activity. Addition of PHA to the postnuclear supernatant of cell lysates also resulted in elevation of phosphatase activity. Elevation of enzyme activity resulted from an increase in the amount of antigen in the immunoprecipitates. Elevation of the quantity was not the result of a de novo protein synthesis since the presence of cycloheximide, a protein synthesis inhibitor, did not modulate the effect of PHA. The effect of PHA was specific since ConA, that also bound to the CD45 molecules, or crosslinking of the antigen by antibody did not affect CD45. Since direct binding of PHA to CD45 molecules was shown in immunoblotting analysis, we suggest that the effect of PHA is a consequence of a PHA-induced conformational change of CD45 that results in up-regulation of the analyzed CD45 epitopes.