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Globally, >100 million cases and >2.6 million deaths had been attributed to coronavirus disease (COVID-19) as of March 14, 2021 (2). Most cases have been reported in Europe and the [...]

Background The clinical and microbial factors associated with Klebsiella pneumoniae bloodstream infections (BSIs) are not well characterized. Prior studies have focused on highly resistant or hypervirulent isolates, limiting our understanding of K. pneumoniae strains that commonly cause BSI. We performed a record review and whole-genome sequencing to investigate the clinical characteristics, bacterial diversity, determinants of antimicrobial resistance, and risk factors for in-hospital death in a cohort of patients with K. pneumoniae BSI. Methods We identified 562 patients at Massachusetts General Hospital with K. pneumoniae BSIs between 2016 and 2022. We collected data on comorbid conditions, infection source, clinical outcomes, and antibiotic resistance and performed whole-genome sequencing on 108 sequential BSI isolates from 2021 to 2022. Results Intra-abdominal infection was the most common source of infection accounting for 34% of all BSIs. A respiratory tract source accounted for 6% of BSIs but was associated with a higher in-hospital mortality rate (adjusted odds ratio, 5.4 [95% confidence interval, 2.2–12.8]; P <.001 for comparison with other sources). Resistance to the first antibiotic prescribed was also associated with a higher risk of death (adjusted odds ratio, 5.2 [95% confidence interval, 2.2–12.4]; P <.001). BSI isolates were genetically diverse, and no clusters of epidemiologically and genetically linked cases were observed. Virulence factors associated with invasiveness were observed at a low prevalence, although an unexpected association between O-antigen type and the source of infection was found. Conclusions These observations demonstrate the versatility of K. pneumoniae as an opportunistic pathogen and highlight the need for new approaches for surveillance and the rapid identification of patients with invasive antimicrobial-resistant K. pneumoniae infection. [ABSTRACT FROM AUTHOR]

We report on the sensitivity and specificity of stool culture compared to polymerase chain reaction for detecting Vibrio cholerae in Haiti during the waning period of the initial outbreak in 2018–2019. We found that stool culture (with a sensitivity of 33.3% and specificity of 97.4%) may not be sufficiently robust in this context. [ABSTRACT FROM AUTHOR]

Relatively few coronavirus disease cases and deaths have been reported from sub-Saharan Africa, although the extent of its spread remains unclear. During August 10-September 11, 2020, we recruited 2,214 participants for a representative household-based cross-sectional serosurvey in Juba, South Sudan. We found 22.3% of participants had severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) receptor binding domain IgG titers above prepandemic levels. After accounting for waning antibody levels, age, and sex, we estimated that 38.3% (95% credible interval 31.8%-46.5%) of the population had been infected with SARS-CoV-2. At this rate, for each PCR-confirmed SARS-CoV-2 infection reported by the Ministry of Health, 103 (95% credible interval 86-126) infections would have been unreported, meaning SARS-CoV-2 has likely spread extensively within Juba. We also found differences in background reactivity in Juba compared with Boston, Massachusetts, USA, where the immunoassay was validated. Our findings underscore the need to validate serologic tests in sub-Saharan Africa populations. [ABSTRACT FROM AUTHOR]

We measured plasma and/or serum antibody responses to the receptor-binding domain (RBD) of the spike (S) protein of SARS-CoV-2 in 343 North American patients infected with SARS-CoV-2 (of which 93% required hospitalization) up to 122 days after symptom onset and compared them to responses in 1548 individuals whose blood samples were obtained prior to the pandemic. After setting seropositivity thresholds for perfect specificity (100%), we estimated sensitivities of 95% for IgG, 90% for IgA, and 81% for IgM for detecting infected individuals between 15 and 28 days after symptom onset. While the median time to seroconversion was nearly 12 days across all three isotypes tested, IgA and IgM antibodies against RBD were short-lived with median times to seroreversion of 71 and 49 days after symptom onset. In contrast, anti-RBD IgG responses decayed slowly through 90 days with only 3 seropositive individuals seroreverting within this time period. IgG antibodies to SARS-CoV-2 RBD were strongly correlated with anti-S neutralizing antibody titers, which demonstrated little to no decrease over 75 days since symptom onset. We observed no cross-reactivity of the SARS-CoV-2 RBD-targeted antibodies with other widely circulating coronaviruses (HKU1, 229 E, OC43, NL63). These data suggest that RBD-targeted antibodies are excellent markers of previous and recent infection, that differential isotype measurements can help distinguish between recent and older infections, and that IgG responses persist over the first few months after infection and are highly correlated with neutralizing antibodies. [ABSTRACT FROM AUTHOR]

Previously we identified an intra-S-phase cell cycle checkpoint elicited by the DNA-damaging carcinogen benzo[α]pyrene-dihydrodiol epoxide (BPDE). Here we have investigated the roles of lesion bypass DNA polymerases polκ and polΒ in the BPDE-induced S-phase checkpoint. BPDE treatment induced the re-localization of an ectopically expressed green fluorescent proteinpolκ fusion protein to nuclear foci containing sites of active DNA synthesis in human lung carcinoma H1299 cells. In contrast, a similarly expressed yellow fluorescent protein-pohΒ fusion protein showed a constitutive nuclear focal distribution at replication forks (in the same cells) that was unchanged in response to BPDE. BPDE-induced formation of green fluorescent protein-polκ nuclear foci was temporally coincident with checkpoint-mediated S-phase arrest. Unlike ‘wild-type’ cells, Polk-/- mouse embryonic fibroblasts (MEFs) failed to recover from BPDE-induced S-phase arrest, while exhibiting normal recovery from S-phase arrest induced by ionizing radiation and hydroxyurea. XPV fibroblasts lacking polΒ showed a normal S-phase checkpoint response to BPDE (but failed to recover from the UV light-induced S-phase checkpoint), in sharp contrast to Polk-/- MEFs. The persistent S-phase arrest in BPDE-treated Polk-/- cells was associated with increased levels of histone γH2AX (a marker of DNA double-strand breaks (DSBs)) and activation of the DSB-responsive kinases ATM and Chk2. These data suggest that in the absence of polκ, replication forks stall at sites of damage and collapse and generate DSBs. Therefore, we conclude that the trans-lesion synthesis enzyme polκ is specifically required for normal recovery from the BPDE-induced S-phase checkpoint. [ABSTRACT FROM AUTHOR]

We have investigated mechanisms that recruit the translesion synthesis (TLS) DNA polymerase Pol κ to stalled replication forks. The DNA polymerase processivity factor PCNA is monoubiquitinated and interacts with Pol κ in cells treated with the bulky adduct-forming genotoxin benzo[a]pyrene dihydrodiol epoxide (BPDE). A monoubiquitination-defective mutant form of PCNA fails to interact with Pol κ. Small interfering RNA-mediated downregulation of the E3 ligase Rad18 inhibits BPDE-induced PCNA ubiquitination and association between PCNA and Pol κ. Conversely, overexpressed Rad18 induces PCNA ubiquitination and association between PCNA and Pol κ in a DNA damage-independent manner. Therefore, association of Pol κ with PCNA is regulated by Rad18-mediated PCNA ubiquitination. Cells from Rad18-/- transgenic mice show defective recovery from BPDE-induced S-phase checkpoints. In Rad18-/- cells, BPDE induces elevated and persistent activation of checkpoint kinases, indicating persistently stalled forks due to defective TLS. Rad18-deficient cells show reduced viability after BPDE challenge compared with wild-type cells (but survival after hydroxyurea or ionizing radiation treatment is unaffected by Rad18 deficiency). Inhibition of RPA/ATR/Chk1-mediated S-phase checkpoint signaling partially inhibited BPDE-induced PCNA ubiquitination and prevented interactions between PCNA and Pol κ. Taken together, our results indicate that ATR/Chk1 signaling is required for Rad18-mediated PCNA monoubiquitination. Recruitment of Pol κ to ubiquitinated PCNA enables lesion bypass and eliminates stalled forks, thereby attenuating the S-phase checkpoint. [ABSTRACT FROM AUTHOR]

The article discusses research on the possible origins of the 2022 resurgence of cholera in Haiti. Topics explored include the genomic and phenotypic analysis of a Vibrio cholerae strain isolated from a stool sample obtained from a cholera patient, the similarity observed in the phenotypes of the 2010 and 2022 bacteria strains, and the factors which may have contributed to the 2022 outbreak such as waning population immunity and poor sanitation.

Relatively few coronavirus disease cases and deaths have been reported from sub-Saharan Africa, although the extent of its spread remains unclear. During August 10-September 11, 2020, we recruited 2,214 participants for a representative household-based cross-sectional serosurvey in Juba, South Sudan. We found 22.3% of participants had severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) receptor binding domain IgG titers above prepandemic levels. After accounting for waning antibody levels, age, and sex, we estimated that 38.3% (95% credible interval 31.8%-46.5%) of the population had been infected with SARS-CoV-2. At this rate, for each PCR-confirmed SARS-CoV-2 infection reported by the Ministry of Health, 103 (95% credible interval 86-126) infections would have been unreported, meaning SARS-CoV-2 has likely spread extensively within Juba. We also found differences in background reactivity in Juba compared with Boston, Massachusetts, USA, where the immunoassay was validated. Our findings underscore the need to validate serologic tests in sub-Saharan Africa populations. [ABSTRACT FROM AUTHOR]

DNA damage induced by the carcinogen benzo[a]pyrene dihydrodiol epoxide (BPDE) induces a Chk1-dependent S-phase checkpoint. Here, we have investigated the molecular basis of BPDE-induced S-phase arrest. Chk1-dependent inhibition of DNA synthesis in BPDE-treated cells occurred without detectable changes in Cdc25A levels, Cdk2 activity, or Cdc7/Dbf4 interaction. Overexpression studies showed that Cdc25A, cyclin A/Cdk2, and Cdc7/Dbf4 were not rate-limiting for DNA synthesis when the BPDE-induced S-phase checkpoint was active. To investigate other potential targets of the S-phase checkpoint, we tested the effects of BPDE on the chromatin association of DNA replication factors. The levels of chromatin- associated Cdc45 (but not soluble Cdc45) were reduced concomitantly with BPDE-induced Chk1 activation and inhibition of DNA synthesis. The chromatin association of Mcm7, Mcm10, and proliferating cell nuclear antigen was unaffected by BPDE treatment. However, the association between Mcm7 and Cdc45 in the chromatin fraction was inhibited in BPDE-treated cells. Chromatin immunoprecipitation analyses demonstrated reduced association of Cdc45 with the β-globin origin of replication in BPDE-treated cells. The inhibitory effects of BPDE on DNA synthesis, Cdc45/Mcm7 associations, and interactions between Cdc45 and the β-globin locus were abrogated by the Chkl inhibitor UCN-O1. Taken together, our results show that the association between Cdc45 and Mcm7 at origins of replication is negatively regulated by Chk1 in a Cdk2-independent manner, Therefore, Cdc45 is likely to be an important target of the Chk1-mediated S-phase checkpoint. [ABSTRACT FROM AUTHOR]