Use of antibodies to the lipopolysaccharide core region in the treatment of gram-negative sepsis and shock

In this paper we review evidence in favor of a protective role for antibodies directed at the lipopolysaccharide (LPS) core region of Gram-negative bacilli. It will be shown that, given the right animal model, polyclonal antisera raised against O-antigen lacking, rough mutant strains, can be shown to protect animals against lethal sepsis due to a variety a bacterial species. Evidence for a protective role obtained from clinical studies will also be discussed. It will be stressed how difficult it is to prove that the antibodies directed against the LPS-core, and not other proteins present in the polyclonal anti-rough strain antisera, were responsible for the observed protective effects. The solution to some of these problems is the production of monoclonal antibodies (Mabs). We will discuss the findings that purified anti-core Mabs may give false-positive outcomes of protection studies, due to the induction of tolerance by small amounts of contaminating endotoxin, present in the antibody preparation. The need to administer the Mab therapeutically instead of prophylactically to the test animals, will be stressed. It will be shown how we succeeded in determining the epitope-specificities of several anti-core antibodies. It will be shown also that use of Elisa alone for characterization of anti-core Mabs may lead to false conclusions concerning epitope specificities. We will prove that synthetic KDO-(=ketodeoxycctanate) and lipid A-containing antigens are indispensable tools for thorough characterization of Mabs. Using this methodology we were able to discriminate between specific antigen-antibody interactions, and the ability of some Mabs to bind “non-specifically” to hydrophobic surfaces. The relationship between epitope specificity and protective effects of anti-core Mabs will be described. Finally, we will demonstrate that a KDO-specific Mabs is capable of protecting mice against lethal sepsis; in other words, it will be shown conclusively that Mabs to defined epitopes in LPS core region are cross-protective.     

The catalytic potency of β-glucosidase from Pyrococcus furiosus in the direct glucosylation reaction

Enzymes from extremophiles operate at conditions that are different from their ‘normal’ counterparts, and are therefore a useful extension of the enzyme toolbox. In this paper, the direct glucosylation reaction mediated by a hyperthermophilic β-glucosidase from Pyrocuccus furiosus was investigated. Hexanol was successfully coupled to glucose with this enzyme. A preliminary study was conducted to improve the product yield. A maximum product concentration of 12.9 g.l⁻¹ was attainable by increasing the glucose concentration to the maximum solubility of 2000 g.(kg buffer solution)⁻¹ at the reaction temperature. The highest glucose based yield of 2.64% was achieved with a glucose concentration of 900 g.(kg buffer solution)⁻¹ at a reaction temperature of 65°C and a pH of 6.0. Performing the reaction at higher pH and temperature led to lower product concentrations. This was caused by deactivation of the enzyme accompanied by browning of the reaction mixture. A pH of 4.4 did have a negative effect on both the storage and the operational stability of the enzyme.     

Polarity of meiotic gene conversion is 5′ to 3′ within the niaD gene of Aspergillus nidulans

We have examined polarity of meiotic gene conversion in the niiA-niaD gene cluster of Aspergillus nidulans in two-point crosses. The type and position of the mutations represented by the niaD alleles and the correlation between the relative frequency of gene conversion and the physical position of these mutations were determined. We show that polarity of meiotic gene conversion is 5′ to 3′ (transcribed strand) within the niaD gene. Additional crosses involving a niiA allele and a niaD allele show little polarity of gene conversion, which suggests that the recombination events leading to restoration of the niaD gene are initiated upstream of the coding region of the niaD gene but within the niiA-niaD gene cluster, possibly within the intergenic promoter region.     

PFGE diversity within the methicillin-resistant <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> clonal lineage ST398

Background  

Livestock has recently been identified as a new reservoir of methicillin-resistant Staphylococcus aureus (MRSA). Most isolates belong to ST398 and are non-typeable with PFGE using SmaI, making it difficult to study transmission and outbreaks. Therefore, a new PFGE using Cfr9I, a neoschizomer of SmaI was optimized and evaluated to investigate ST398 isolates.     

Asymptomatic Middle East Respiratory Syndrome Coronavirus Infection in Rabbits

The ability of Middle East respiratory syndrome coronavirus (MERS-CoV) to infect small animal species may be restricted given the fact that mice, ferrets, and hamsters were shown to resist MERS-CoV infection. We inoculated rabbits with MERS-CoV. Although virus was detected in the lungs, neither significant histopathological changes nor clinical symptoms were observed. Infectious virus, however, was excreted from the upper respiratory tract, indicating a potential route of MERS-CoV transmission in some animal species.