Streptococcus pneumoniae G5 domains bind different ligands

Many bacterial pathogens express small G5 domains that exist in the context of various membrane‐anchored proteins and these G5 domains have been associated with colonization, cellular adhesion, and biofilm formation. However, despite over a decade since the computational prediction of these G5 domains, many remain uncharacterized, particularly those from Streptococcus pneumoniae. Of five previously predicted G5 domains we found that four of these, all derived from S. pneumoniae, are independently folded modules. As one of these exhibits extreme line broadening due to self‐association, we were able to use NMR solution studies to probe the potential ligand interactions of the remaining three G5 domains. None of these G5 domains engage N‐acetylglucosamine (NAG) as previously predicted but do interact with other small molecules that may modulate adherence to both bacteria and host cells. Specifically, while all G5 domains tested engage Zn, only one of these G5 domains engage heparin. NMR solution structural studies of the IgA1 Protease G5 (IgA1P‐G5) and endo‐beta‐N‐acetylglucosaminidase‐D G5 (ENDD‐G5) also facilitated identification of the ligand binding sites and confirm the typical G5 fold that comprises two connected β‐sheets with no canonical core. NMR relaxation experiments indicate flexibility on both ends and within the connecting regions between the β‐sheets. Our studies thus establish a basis for future biological experiments to test whether the ligands presented here are involved in bacterial adherence, either to bacteria or to host cells.     

Mother-to-Child HIV-1 Transmission Events Are Differentially Impacted by Breast Milk and Its Components from HIV-1-Infected Women

Breast milk is a vehicle of infection and source of protection in post-natal mother-to-child HIV-1 transmission (MTCT). Understanding the mechanism by which breast milk limits vertical transmission will provide critical insight into the design of preventive and therapeutic approaches to interrupt HIV-1 mucosal transmission. However, characterization of the inhibitory activity of breast milk in human intestinal mucosa, the portal of entry in postnatal MTCT, has been constrained by the limited availability of primary mucosal target cells and tissues to recapitulate mucosal transmission ex vivo. Here, we characterized the impact of skimmed breast milk, breast milk antibodies (Igs) and non-Ig components from HIV-1-infected Ugandan women on the major events of HIV-1 mucosal transmission using primary human intestinal cells and tissues. HIV-1-specific IgG antibodies and non-Ig components in breast milk inhibited the uptake of Ugandan HIV-1 isolates by primary human intestinal epithelial cells, viral replication in and transport of HIV-1- bearing dendritic cells through the human intestinal mucosa. Breast milk HIV-1-specific IgG and IgA, as well as innate factors, blocked the uptake and transport of HIV-1 through intestinal mucosa. Thus, breast milk components have distinct and complementary effects in reducing HIV-1 uptake, transport through and replication in the intestinal mucosa and, therefore, likely contribute to preventing postnatal HIV-1 transmission. Our data suggests that a successful preventive or therapeutic approach would require multiple immune factors acting at multiple steps in the HIV-1 mucosal transmission process.     

Streptococcus pneumoniae IgA1 protease: A metalloprotease that can catalyze in a split manner in vitro

IgA1 proteases (IgA1P) from diverse pathogenic bacteria specifically cleave human immunoglobulin A1 (IgA1) at the hinge region, thereby thwarting protective host immune responses. Streptococcus pneumoniae (S. pneumoniae) IgA1P shares no sequence conservation with serine or cysteine types of IgA1Ps or other known proteins, other than a conserved HExxH Zn‐binding motif (1604‐1608) found in metalloproteases. We have developed a novel expression system to produce the mature S. pneumoniae IgA1P and we have discovered that this form is both attached to the bacterial cell surface and released in its full form. Our data demonstrate that the S. pneumoniae IgA1P comprises two distinct regions that associate to form an active metalloprotease, the first such example of a metalloprotease that can be split in vitro and recombined to form an active enzyme. By capitalizing on this novel domain architecture, we show that the N‐terminal region of S. pneumoniae IgA1P comprises the primary binding region for IgA1, although the C‐terminal region of S. pneumoniae IgA1P is necessary for cleavage of IgA1. Our findings lend insight into the protein domain architecture of the S. pneumoniae IgA1P and function of this important virulence factor for S. pneumoniae infection.     

Evaluation of a pumping system for sampling zooplankton

This study describes the sampling efficiency of a plankton pumpdesigned to operate from a moving research vessel. The samplingefficiency of the pump was evaluated by comparing the catchper unit volume of several planktonic taxa sampled simultaneouslywith the pump and with a modified Gulf V plankton sampler ora tube sampler. The estimates of plankton density obtained withthe pump sampler corresponded well with those obtained withthe modified plankton net and tube sampler. We conclude thatthe use of a plankton pump in zooplankton sampling gives resultscomparable to those found with traditional sampling, exceptthat the results were biased for the sampling of rotifers andcopepod eggs.     

Comparison of methods used in zooplankton sampling and counting in the joint Russian-Finnish evaluation of the trophic state of Lake Ladoga

The results of a joint Russian-Finnish investigation on zooplankton in Lake Ladoga are presented, and a comparison is made between two sampling techniques, tube sampler and plankton net, and between two counting methods. The precision of subsampling and sampling is further discussed on the basis of zooplankton data gathered in Lake Saimaa, Finland. The comparison clearly indicates that a tube sampler is required for reliable sampling of small-sized animals, while a plankton net saves time and is a more economical sampler for large, rare or active animals. The comparison between the results obtained by Finnish and Russian workers, using different counting procedures, shows that the main groups of crustacean zooplankton are similarly counted and identified in the two laboratories.