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AbstractAn outbreak of serotype 19A Streptococcus pneumoniaeoccurred among the residents of a relief facility. Pneumonia developed in 5 of 99 residents (attack rate, 5.1%). We obtained pharyngeal specimens from non-onset residents, and S. pneumoniaewas isolated from 6 individuals (6.4%), 5 of whom had serotype 19A.

ABSTRACTThe treatment regimen for diarrhea depends greatly on correct diagnosis of its etiology. Recent diarrhea outbreaks in Bangladesh showed Vibrio choleraeto be the predominant cause, although more than 40% of the suspected cases failed to show cholera etiology by conventional culture methods (CMs). In the present study, suspected cholera stools collected from every 50th patient during an acute diarrheal outbreak were analyzed extensively using different microbiological and molecular tools to determine their etiology. Of 135 stools tested, 86 (64%) produced V. choleraeO1 by CMs, while 119 (88%) tested positive for V. choleraeO1 by rapid cholera dipstick (DS) assay; all but three samples positive for V. choleraeO1 by CMs were also positive for V. choleraeO1 by DS assay. Of 49 stools that lacked CM-based cholera etiology despite most being positive for V. choleraeO1 by DS assay, 25 (51%) had coccoid V. choleraeO1 cells as confirmed by direct fluorescent antibody (DFA) assay, 36 (73%) amplified primers for the genes wbeO1 and ctxAby multiplex-PCR (M-PCR), and 31 (63%) showed El Tor-specific lytic phage on plaque assay (PA). Each of these methods allowed the cholera etiology to be confirmed for 97% of the stool samples. The results suggest that suspected cholera stools that fail to show etiology by CMs during acute diarrhea outbreaks may be due to the inactivation of V. choleraeby in vivovibriolytic action of the phage and/or nonculturability induced as a host response.

ABSTRACTVibrio choleraeO1 biotype El Tor (ET), the cause of the current 7th pandemic, has recently been replaced in Asia and Africa by an altered ET biotype possessing cholera toxin (CTX) of the classical (CL) biotype that originally caused the first six pandemics before becoming extinct in the 1980s. Until recently, the ET prototype was the biotype circulating in Peru; a detailed understanding of the evolutionary trend of V. choleraecausing endemic cholera in Latin America is lacking. The present retrospective microbiological, molecular, and phylogenetic study of V. choleraeisolates recovered in Mexico (n= 91; 1983 to 1997) shows the existence of the pre-1991 CL biotype and the ET and CL biotypes together with the altered ET biotype in both epidemic and endemic cholera between 1991 and 1997. According to sero- and biotyping data, the altered ET, which has shown predominance in Mexico since 1991, emerged locally from ET and CL progenitors that were found coexisting until 1997. In Latin America, ET and CL variants shared a variable number of phenotypic markers, while the altered ET strains had genes encoding the CL CTX (CTXCL) prophage, ctxBCLand rstRCL, in addition to resident rstRET, as the underlying regional signature. The distinct regional fingerprints for ET in Mexico and Peru and their divergence from ET in Asia and Africa, as confirmed by subclustering patterns in a pulsed-field gel electrophoresis (NotI)-based dendrogram, suggest that the Mexico epidemic in 1991 may have been a local event and not an extension of the epidemics occurring in Asia and South America. Finally, the CL biotype reservoir in Mexico is unprecedented and must have contributed to the changing epidemiology of global cholera in ways that need to be understood.

A real‐time PCR assay with the cycling probe method was used to detect mutations at codons 83 and 87 in the DNA gyrase A subunit encoded by gyrAin Salmonella entericaserovar Typhi and Paratyphi A clinical isolates. The susceptibility estimated from the results of the gyrAmutation assay was consistent with that identified by the culture method using an E‐test. This assay allows rapid screening of S. entericaserovar Typhi and Paratyphi A with reduced susceptibility to ciprofloxacin.

Clinical isolates of Streptococcus pyogeneswere classified by serological typing of their surface M protein. Non‐M typeable strains with the emm1gene were characterized as the degradation of M protein caused by overproduction of the extracellular cysteine protease, SpeB. These events are dependent on the growth phase. M protein produced prior to expression of SpeB is degraded in the stationary phase when the active form of SpeB is detected. The proteolytic degradation of M protein should be considered for precise M typing analysis.

To analyze the antibacterial activity of Bacillus amyloliquefaciensphage endolysin, nine deletion derivatives of the endolysin were constructed. Each deletion mutant was overexpressed, purified and characterized. The catalytic domain was located on the N‐terminal region and the C‐terminus had an affinity with the bacterial envelope. The enzymatic activity remained in spite of the deletion of the C‐terminal 116‐amino acid region; however, the antibacterial activity was lost. These results indicate that antibacterial action requires both the C‐terminal cell‐binding and the N‐terminal enzymatic activities.

Self‐disruptive Escherichia colithat produces foreign target protein was developed. E. coliwas co‐transformed with two vector plasmids, a target gene expression vector and a lysis gene expression vector. The lytic protein was produced after the expression of the target gene, resulting in simplification of the cell disruption process. In this study, the expression of cloned T4 phage gene eor twas used for the disruption of E. colithat produced β‐glucuronidase (GUS) as a model target protein. The expression of gene edid not lead to prompt cell disruption but weakened the cell wall. Resuspension with deionized water facilitated cell lysis, and GUS activity was observed in the resuspended liquid. Expression of gene eat mid logarithmic growth phase was the optimal induction period for GUS production and release. On the other hand, the expression of gene tinduced immediate cell lysis, and intracellular GUS was released to the culture medium. Maximum GUS production was obtained when gene twas induced at late logarithmic growth phase.

To analyze the antibacterial activity of Bacillus amyloliquefaciensphage endolysin, nine deletion derivatives of the endolysin were constructed. Each deletion mutant was overexpressed, purified and characterized. The catalytic domain was located on the N-terminal region and the C-terminus had an affinity with the bacterial envelope. The enzymatic activity remained in spite of the deletion of the C-terminal 116-amino acid region; however, the antibacterial activity was lost. These results indicate that antibacterial action requires both the C-terminal cell-binding and the N-terminal enzymatic activities.

Cholera continues to be a global concern, as large epidemics have occurred recently in Haiti, Yemen, and countries of sub-Saharan Africa. A single lineage of Vibrio choleraeO1 has been considered to be introduced into these regions from South Asia and to cause the spread of cholera. Using genomic epidemiology, we showed that two distinct lineages exist in Bengal, one of which is linked to the global lineage. The other lineage was found only in Iran, Iraq, and countries in Asia and differed from the global lineage regarding cholera toxin variant and drug resistance profile. Therefore, the potential transmission of this lineage to other regions would likely cause worldwide cholera spread and may result in this lineage replacing the current global lineage.

Antimicrobial peptides play an important role in host defense against Vibrio cholerae. Generally, the V. choleraeO1 classical biotype is polymyxin B (PB) sensitive and El Tor is relatively resistant. Detection of classical biotype traits like the production of classical cholera toxin and PB sensitivity in El Tor strains has been reported in recent years, including in the devastating Yemen cholera outbreak during 2016-2018. To investigate the factor(s) responsible for the shift in the trend of sensitivity to PB, we studied the two-component system encoded by carRS, regulating the lipid A modification of El Tor vibrios, and found that only carRcontains a single nucleotide polymorphism (SNP) in recently emerged PB-sensitive strains.

ABSTRACTStreptococcus pneumoniaecolonizes the nasopharyngeal mucus in healthy individuals and can cause otitis media, pneumonia, and invasive pneumococcal diseases. In this study, we analyzed S. pneumoniaestrains that caused 19 pneumonia episodes in long-term inpatients with severe underlying disease in a hospital during a period of 14 months (from January 2014 to February 2015). Serotyping and whole-genome sequencing analyses revealed that 18 of the 19 pneumonia cases were caused by S. pneumoniaestrains belonging to 3 genetically distinct groups: clonal complex 9999 (CC9999), sequence type 282 (ST282), and ST166. The CC9999 and ST282 strains appeared to have emerged separately by a capsule switch from the pandemic PMEN 1 strain (Spain23F-ST81). After all the long-term inpatients were inoculated with the 23-valent pneumococcal polysaccharide vaccine, no other nosocomial pneumonia infections occurred until March 2016.

ABSTRACTThis study characterized a cephalosporin-resistant Salmonella entericaserovar Typhi isolate. The organism possessed a plasmid encoding the CTX-M-15 extended-spectrum-β-lactamase. This plasmid is the determinant for the phenotype of cephalosporin resistance and is transferrable among Enterobacteriaceae.