Staphylococcus aureus Strain USA300 Perturbs Acquisition of Lysosomal Enzymes and Requires Phagosomal Acidification for Survival inside Macrophages

Abstract

Methicillin-resistant Staphylococcus aureus (MRSA) causes invasive, drug-resistant skin and soft tissue infections. Reports that S. aureus bacteria survive inside macrophages suggest that the intramacrophage environment may be a niche for persistent infection; however, mechanisms by which the bacteria might evade macrophage phagosomal defenses are unclear. We examined the fate of the S. aureus-containing phagosome in THP-1 macrophages by evaluating bacterial intracellular survival and phagosomal acidification and maturation and by testing the impact of phagosomal conditions on bacterial viability. Multiple strains of S. aureus survived inside macrophages, and in studies using the MRSA USA300 clone, the USA300-containing phagosome acidified rapidly and acquired the late endosome and lysosome protein LAMP1. However, fewer phagosomes containing live USA300 bacteria than those containing dead bacteria associated with the lysosomal hydrolases cathepsin D and ␤-glucuronidase. Inhibiting lysosomal hydrolase activity had no impact on intracellular survival of USA300 or other S. aureus strains, suggesting that S. aureus perturbs acquisition of lysosomal enzymes. We examined the impact of acidification on S. aureus intramacrophage viability and found that inhibitors of phagosomal acidification significantly impaired USA300 intracellular survival. Inhibition of macrophage phagosomal acidification resulted in a 30-fold reduction in USA300 expression of the staphylococcal virulence regulator agr but had little effect on expression of sarA, saeR, or sigB. Bacterial exposure to acidic pH in vitro increased agr expression. Together, these results suggest that S. aureus survives inside macrophages by perturbing normal phagolysosome formation and that USA300 may sense phagosomal conditions and upregulate expression of a key virulence regulator that enables its intracellular survival. Staphylococcus aureus is the primary cause of skin and soft tissue infections (SSTIs) in humans. In the United States alone, approximately 14 million people seek medical treatment each year for SSTIs associated with methicillin-sensitive or methicillin-resistant S. aureus (MSSA or MRSA, respectively) (1), and over 50% of cases are caused by MRSA strains (2). MRSA infections can persist and disseminate to deeper sites in the host, causing diseases such as endocarditis, osteomyelitis, or bacteremia, and are estimated to cause over 18,000 deaths per year in the United States (3). High hospitalization and mortality rates associated with MRSA are attributed to the bacterium's increasing drug resistance; MRSA strains are resistant to the beta-lactam drugs penicillin, methicillin, and oxacillin, and the emergence of vancomycin-resistant strains (4) means that few treatment options remain.S. aureus was once recognized primarily as a hospital-acquired (HA) pathogen that gained access to the host via indwelling medical devices. However, strains of community-acquired MRSA (CA-MRSA) have emerged that infect healthy individuals with no predisposing risk factors f...

Identification and Characterization of Staphylococcus aureus Strains with an Incomplete Hemolytic Phenotype

“…PCR testing found that all 60 SIHP strains possessed the tst gene suggesting increased virulence. Recently, many reports showed that S. aureus could survive in the human monocyte-derived macrophages and these S. aureus strains are more virulent (Kubica et al, 2008; Tranchemontagne et al, 2015; Münzenmayer et al, 2016; Nandi and Bishayi, 2016). Therefore, we compared the intracellular survival abilities of SIHP strains and SCHP strains which is tst negative S. aureus in macrophages, and found that intracellular survival ability of SIHP strain was much higher than that of tst negative S. aureus strain.…”

In or out: Phagosomal escape ofStaphylococcus aureus

“…SaCP maturation quickly acquires the late phagosomal marker lysosome‐associated membrane protein‐1 (LAMP‐1) and Rab7. LAMP‐1 positivity and acidification of SaCP seem to be independent of host species and cell type (see, for instance, Kubica et al, ; Giese et al, ; Lâm et al, ; Grosz et al, ; Flannagan, Heit, & Heinrichs, ; Tranchemontagne, Camire, O'Donnell, Baugh, & Burkholder, ; Jubrail et al, ; Lacoma et al, ) indicating that S . aureus does not interfere with initial phagosomal maturation.…”

“…Although other lysosomal markers such as LAMP‐2 (Flannagan et al, ; Flannagan et al, ) have been identified in SaCP, some studies suggest the absence of a fully functional phagolysosomal microenvironment of internalised S . aureus because key lysosomal hydrolases such as cathepsin D and beta‐glucuronidase are not recruited to the SaCP (Jubrail et al, ; Tranchemontagne et al, ), and SaCP fails to acidify and acquire LIMP‐2 (Jubrail et al, ).…”

Staphylococcus aureus intracellular survival: A closer look in the process

“…Recent report from Tranchemontagne et al 7 show that S aureus survived inside macrophage by inhibiting normal phagolysosomal formation causing its intracellular survival. Another study from Kulkarni et al 8 show that in response to exposure of cigarette smoke extract, S aureus was able to supress host immune response and also modulate several virulence genes which help its evasion from bactericidal function of leukocytes.…”

“…Previously it was reported that S aureus favors an acidic environment and this strategy help intracellular survival. 7 Authors also investigated the contribution of acidic environment on S aureus survival in this model using lysotracker and found that acidification do not play a role in this mechanism.…”