Streptococcus pneumoniae is the most common causative agent of community-acquired pneumonia and can penetrate epithelial barriers to enter the bloodstream and brain. We investigated intracellular fates of S. pneumoniae and found that the pathogen is entrapped by selective autophagy in pneumolysin- and ubiquitin-p62-LC3 cargo-dependent manners. Importantly, following induction of autophagy, Rab41 was relocated from the Golgi apparatus to S. pneumoniae-containing autophagic vesicles (PcAV), which were only formed in the presence of Rab41-positive intact Golgi apparatuses. Moreover, subsequent localization and regulation of K48- and K63-linked polyubiquitin chains in and on PcAV were clearly distinguishable from each other. Finally, we found that E3 ligase Nedd4-1 was recruited to PcAV and played a pivotal role in K63-linked polyubiquitin chain (K63Ub) generation on PcAV, promotion of PcAV formation, and elimination of intracellular S. pneumoniae. These findings suggest that Nedd4-1-mediated K63Ub deposition on PcAV acts as a scaffold for PcAV biogenesis and efficient elimination of host cell-invaded pneumococci.

ABSTRACTLegionella pneumophilaserogroup (SG) 1 is the most frequent cause of legionellosis. This study analyzed environmental isolates ofL. pneumophilaSG 1 in Japan using monoclonal antibody (MAb) typing and sequence-based typing (SBT). Samples were analyzed from bathwater (BW;n= 50), cooling tower water (CT;n= 50), and soil (SO;n= 35). The distribution of MAb types varied by source, with the most prevalent types being Bellingham (42%), Oxford (72%), and OLDA (51%) in BW, CT, and SO, respectively. The ratios of MAb 3/1 positive isolates were 26, 2, and 14% from BW, CT, and SO, respectively. The environmental isolates from BW, CT, and SO were divided into 34 sequence types (STs; index of discrimination [IOD] = 0.973), 8 STs (IOD = 0.448), and 11 STs (IOD = 0.879), respectively. Genetic variation among CT isolates was smaller than seen in BW and SO. ST1 accounted for 74% of the CT isolates. The only common STs between (i) BW and CT, (ii) BW and SO, and (iii) CT and SO were ST1, ST129, and ST48, respectively, suggesting that each environment constitutes an independent habitat.

The Reference Center in Japan collected 427 clinical isolates between 2008 and 2016, including 7 representative isolates from corresponding outbreaks. The collection included 419 isolates, of which 372 belonged to serogroup 1 (SG1) (87%) and the others belonged to SG2 to SG15 except for SG7 and SG11, and 8 isolates of other species (, ,, ,, and ). isolates were genotyped by sequence-based typing (SBT) and represented 187 sequence types (STs), of which 126 occurred in a single isolate (index of discrimination of 0.984). These STs were analyzed using minimum spanning tree analysis, resulting in the formation of 18 groups. The pattern of overall ST distribution among isolates was diverse. In particular, some STs were frequently isolated and were suggested to be related to the infection sources. The major STs were ST23 (35 isolates), ST120 (20 isolates), and ST138 (16 isolates). ST23 was the most prevalent and most causative ST for outbreaks in Japan and Europe. ST138 has been observed only in Japan, where it has caused small-scale outbreaks; 81% of those strains (13 isolates) were suspected or confirmed to infect humans through bath water sources. On the other hand, 11 ST23 strains (31%) and 5 ST120 strains (25%) were suspected or confirmed to infect humans through bath water. These findings suggest that some ST strains frequently cause legionellosis in Japan and are found under different environmental conditions. serogroup 1 (SG1) is the most frequent cause of legionellosis. Our previous genetic analysis indicated that SG1 environmental isolates represented 8 major clonal complexes, consisting of 3 B groups, 2 C groups, and 3 S groups, which included major environmental isolates derived from bath water, cooling towers, and soil and puddles, respectively. Here, we surveyed clinical isolates collected from patients with legionellosis in Japan between 2008 and 2016. Most strains belonging to the B group were isolated from patients for whom bath water was the suspected or confirmed source of infection. Among the isolates derived from patients whose suspected infection source was soil or dust, most belonged to the S1 group and none belonged to the B or C groups. Additionally, the U group was discovered as a new group, which mainly included clinical isolates with unknown infection sources.

A conspicuous adhesion of Staphylococcus aureus organisms to murine cutaneous fibroblasts and NIH/ 3T3 cells cultured in vitro and subsequent ingestion of S. aureus organisms by these fibroblasts are described.In the present experimental system, only fibroblasts-adhering S. aureus organisms were efficiently ingested by fibroblasts unlike S. epidermidis and S. saprophyticus. These findings might suggest a correlation between the pathogenesis of S. aureus and its intracellular localization in non-professional phagocytes such as fibroblasts in a special reference to its higher pathogenicity than those of coagulase negative counterparts.

The virulence of a pleiotropic Staphylococcus aureus mutant with an extremely low producibility of coagulase and other factors was investigated in mouse. A mutant strain, designated as CL-1, showed the same LD50 and the same intrarenal proliferation as its parental strain, when the mutant organisms were inoculated in mice in high doses. The mutant organisms, however, showed a diminished intrarenal proliferation compared with its parental organisms in low doses. This mutant strain expressed a pleiotropic phenotype such as a concomitant reduction in the producibility of coagulase, a-toxin, and Panton-Valentine leucocidin. The total effect due to the reduction in producibility of various factors on the virulence of the mutant strain was investigated with studies on the bacterial resistance to the phagocytic activity of leucocytes.A possible role of coagulase and that of some other staphylococcal exoproteins in the pathogenesis of S. aureus were discussed.