The Haemophilus influenzae Type b hcsA and hcsB Gene Products Facilitate Transport of Capsular Polysaccharide across the Outer Membrane and Are Essential for Virulence

Abstract

Haemophilus influenzae type b is a common cause of invasive bacterial disease, especially among children in underdeveloped countries. The type b polysaccharide capsule is a polymer of ribose and ribitol-5-phosphate and is a critical determinant of virulence. Expression of the type b capsule is dependent upon the cap b locus, which consists of three functionally distinct regions, designated regions 1 to 3. Region 3 contains the hcsA and hcsB genes, which share significant homology with genes that have been implicated in encapsulation in other pathogenic bacteria but have unclear functions. In this study, we inactivated hcsA alone, hcsB alone, and both hcsA and hcsB together and examined the effects of these mutations on polysaccharide transport and bacterial virulence properties. Inactivation of hcsA alone resulted in accumulation of polysaccharide in the periplasm and a partial decrease in surface-associated polysaccharide, whereas inactivation of hcsB alone or of both hcsA and hcsB together resulted in accumulation of polysaccharide in the periplasm and complete loss of surfaceassociated polysaccharide. All mutations eliminated serum resistance and abrogated bacteremia and mortality in neonatal rats. These results indicate that the hcsA and hcsB gene products have complementary functions involved in the transport of polysaccharide across the outer membrane and are essential for virulence.

KpsC and KpsS are retaining 3-deoxy- d - manno -oct-2-ulosonic acid (Kdo) transferases involved in synthesis of bacterial capsules

“…KpsC and KpsS are known to be integral parts of a multiprotein CPS assembly complex that contains export machinery (9,29). In H. influenzae group b, hcsAB mutants are reported to accumulate polymer in the periplasm, leading to the suggestion that the proteins facilitate transport through the outer membrane (31). However, their sequences predict them to be cytoplasmic proteins, and this location has been confirmed experimentally for the E. coli homologs (1,9).…”

Extraction and Purification of Haemophilus influenzae Type b Lipooligosac‌charide by Modified Phenol Method

“…Haemophilus influenzae is a Gram-negative, non-motile and fastidious bacterium classified into two groups of capsular and non-capsular strains [1]. The encapsulated strains are serologically subdivided into 6 (a -f) distinct types based on the capsular antigen.…”

Use of bexB To Detect the Capsule Locus in Haemophilus influenzae

“…1B). The genes contained within regions I and III, designated bexDCBA and hcsAB, respectively, are highly conserved across all capsular types and are required for transport of capsule constituents across the outer membrane (20,21,23,37,40). Region II genes encode capsule type a-through f-specific proteins and thus vary by serotype.…”