A bioinformatic approach to identify confirmed and probable CRISPR-Cas systems in the Acinetobacter calcoaceticus-Acinetobacter baumannii complex genomes.

Introduction: The Acinetobacter calcoaceticus-Acinetobacter baumannii complex, or Acb complex, consists of six species: Acinetobacter baumannii, Acinetobacter calcoaceticus, Acinetobacter nosocomialis, Acinetobacter pittii, Acinetobacter seifertii, and Acinetobacter lactucae. A. baumannii is the most clinically significant of these species and is frequently related to healthcare-associated infections (HCAIs). Clustered regularly interspaced short palindromic repeat (CRISPR) arrays and associated genes (cas) constitute bacterial adaptive immune systems and function as variable genetic elements. This study aimed to conduct a genomic analysis of Acb complex genomes available in databases to describe and characterize CRISPR systems and cas genes. Methods: Acb complex genomes available in the NCBI and BV-BRC databases, the identification and characterization of CRISPR-Cas systems were performed using CRISPRCasFinder, CRISPRminer, and CRISPRDetect. Sequence types (STs) were determined using the Oxford scheme and ribosomal multilocus sequence typing (rMLST). Prophages were identified using PHASTER and Prophage Hunter. Results: A total of 293 genomes representing six Acb species exhibited CRISPR-related sequences. These genomes originate from various sources, including clinical specimens, animals, medical devices, and environmental samples. Sequence typing identified 145 ribosomal multilocus sequence types (rSTs). CRISPR-Cas systems were confirmed in 26.3% of the genomes, classified as subtypes I-Fa, I-Fb and I-Fv. Probable CRISPR arrays and cas genes associated with CRISPR-Cas subtypes III-A, I-B, and III-B were also detected. Some of the CRISPR-Cas systems are associated with genomic regions related to Cap4 proteins, and toxin-antitoxin systems. Moreover, prophage sequences were prevalent in 68.9% of the genomes. Analysis revealed a connection between these prophages and CRISPR-Cas systems, indicating an ongoing arms race between the bacteria and their bacteriophages. Furthermore, proteins associated with anti-CRISPR systems, such as AcrF11 and AcrF7, were identified in the A. baumannii and A. pittii genomes. Discussion: This study elucidates CRISPR-Cas systems and defense mechanisms within the Acb complex, highlighting their diverse distribution and interactions with prophages and other genetic elements. This study also provides valuable insights into the evolution and adaptation of these microorganisms in various environments and clinical settings.

Characterization of Cronobacter sakazakii and Cronobacter malonaticus Strains Isolated from Powdered Dairy Products Intended for Consumption by Adults and Older Adults.

The objective of this study was to characterize Cronobacter spp. and related organisms isolated from powder dairy products intended for consumption by adults and older adults using whole-genome sequencing (WGS), and to identify genes and traits that encode antibiotic resistance and virulence. Virulence (VGs) and antibiotic resistance genes (ARGs) were detected with the Comprehensive Antibiotic Resistance Database (CARD) platform, ResFinder, and MOB-suite tools. Susceptibility testing was performed using disk diffusion. Five presumptive strains of Cronobacter spp. were identified by MALDI-TOF MS and ribosomal MLST. Three C. sakazakii strains were of the clinical pathovar ST1, one was ST31, and the remaining isolate was C. malonaticus ST60. In addition, Franconibacter helveticus ST345 was identified. The C. sakazakii ST1 strains were further distinguished using core genome MLST based on 2831 loci. Moreover, 100% of the strains were resistant to cefalotin, 75% to ampicillin, and 50% to amikacin. The C. sakazakii ST1 strains were multiresistant (MDR) to four antibiotics. Additionally, all the strains adhered to the N1E-115 cell line, and two invaded it. Eighteen ARGs mainly involved in antibiotic target alteration and antibiotic efflux were detected. Thirty VGs were detected and clustered as flagellar proteins, outer membrane proteins, chemotaxis, hemolysins, and genes involved in metabolism and stress. The pESA3, pSP291-1, and pCMA1 plasmids were detected, and the prevalent mobile genetic elements (MGEs) were ISEsa1, ISEc52, and IS26. The isolates of C. sakazakii and C. malonaticus exhibited multiresistance to antibiotics, harbored genes encoding various antibiotic resistance proteins, and various virulence factors. Consequently, these contaminated powdered dairy products pose a risk to the health of hypersensitive adults.

Tet2 regulates Sin3a recruitment at active enhancers in embryonic stem cells.

Tet2 is a member of the Ten-eleven translocation (Tet1/2/3) family of enzymes and is expressed in embryonic stem cells (ESCs). It demethylates DNA (catalytic functions) and partners with chromatin modifiers (noncatalytic functions) to regulate genes. However, the significance of these functions in ESCs is less defined. Using Tet2 catalytic mutant (Tet2m/m) and knockout (Tet2-/-) ESCs, we identified Tet2 target genes regulated by its catalytic dependent versus independent roles. Tet2 was enriched at their active enhancers and promoters to demethylate them. We also identified the histone deacetylase component Sin3a as a Tet2 partner, co-localizing at promoters and active enhancers. Tet2 deficiency diminished Sin3a at these regions. Tet2 and Sin3a co-occupancy overlapped with Tet1. Combined loss of Tet1/2, but not of their catalytic activities, reduced Sin3a at active enhancers. These findings establish Tet2 catalytic and noncatalytic functions as regulators of DNA demethylation and Sin3a recruitment at active enhancers in ESCs.

Screening of Antibiotic and Virulence Genes from Whole Genome Sequenced Cronobacter sakazakii Isolated from Food and Milk-Producing Environments.

The objective of this study was to use whole-genome sequencing (WGS) to screen for genes encoding for antibiotic resistance, fitness and virulence in Cronobacter sakazakii strains that had been isolated from food and powdered-milk-producing environments. Virulence (VGs) and antibiotic-resistance genes (ARGs) were detected with the Comprehensive Antibiotic Resistance Database (CARD) platform, ResFinder and PlasmidFinder tools. Susceptibility testing was performed using disk diffusion. Fifteen presumptive strains of Cronobacter spp. were identified by MALDI-TOF MS and ribosomal-MLST. Nine C. sakazakii strains were found in the meningitic pathovar ST4: two were ST83 and one was ST1. The C. sakazakii ST4 strains were further distinguished using core genome MLST based on 3678 loci. Almost all (93%) strains were resistant to cephalotin and 33% were resistant to ampicillin. In addition, 20 ARGs, mainly involved in regulatory and efflux antibiotics, were detected. Ninety-nine VGs were detected that encoded for OmpA, siderophores and genes involved in metabolism and stress. The IncFIB (pCTU3) plasmid was detected, and the prevalent mobile genetic elements (MGEs) were ISEsa1, ISEc52 and ISEhe3. The C. sakazakii isolates analyzed in this study harbored ARGs and VGs, which could have contributed to their persistence in powdered-milk-producing environments, and increase the risk of infection in susceptible population groups.

Comparative analysis of Tet2 catalytic-deficient and knockout bone marrow over time.

TET2 is a member of the Ten-eleven translocation (Tet) family of DNA dioxygenases that regulate gene expression by promoting DNA demethylation (enzymatic activity) and partnering with chromatin regulatory complexes (nonenzymatic functions). TET2 is highly expressed in the hematopoietic lineage, where its molecular functions are the subject of continuous investigations because of the prevalence of TET2 mutations in hematologic malignancies. Previously, we have implicated Tet2 catalytic and noncatalytic functions in the regulation of myeloid and lymphoid lineages, respectively. However, the impact of these functions of Tet2 on hematopoiesis as the bone marrow ages remains unclear. Here, we conducted comparative transplantations and transcriptomic analyses of 3-, 6-, 9-, and 12-month-old Tet2 catalytic mutant (Mut) and knockout (KO) bone marrow. Tet2 Mut bone marrow of all ages exclusively caused hematopoietic disorders of the myeloid lineage. In contrast, young Tet2 KO bone marrow developed both lymphoid and myeloid diseases, whereas older Tet2 KO bone marrow predominantly elicited myeloid disorders with shorter latency than age-matched Tet2 Mut bone marrow. We identified robust gene dysregulation in Tet2 KO Lin- cells at 6 months that involved lymphoma and myelodysplastic syndrome and/or leukemia-causing genes, many of which were hypermethylated early in life. There was a shift from lymphoid to myeloid gene deregulation in Tet2 KO Lin- cells with age, underpinning the higher incidence of myeloid diseases. These findings expand on the dynamic regulation of bone marrow by Tet2 and show that its catalytic-dependent and -independent roles have distinct impacts on myeloid and lymphoid lineages with age.

Local intensity of artisanal gold mining drives mercury accumulation in neotropical oxbow lake fishes.

Driven by surges in global gold prices and additional socio-economic factors, artisanal small-scale gold mining (ASGM) in the Global South is increasing and driving emissions of significant quantities of mercury (Hg) into the air and freshwater. Hg can be toxic to animal and human populations and exacerbate the degradation of neotropical freshwater ecosystems. We examined drivers of Hg accumulation in fish that inhabit oxbow lakes of Peru's Madre de Dios, a region with high biodiversity value and increasing human populations that depend on ASGM. We hypothesized that fish Hg levels would be driven by local ASGM activities, by environmental Hg exposure, by water quality, and by fish trophic level. We sampled fish in 20 oxbow lakes spanning protected areas and areas subject to ASGM during the dry season. Consistent with previous findings, Hg levels were positively associated with ASGM activities, and were higher in larger, carnivorous fish and where water had lower dissolved oxygen levels. In addition, we found a negative relationship between fish mercury levels associated with ASGM and the occurrence of the piscivorous giant otter. The link between fine-scale quantification of spatial ASGM activity and Hg accumulation, as indicated by the result that in the lotic environment, localized effects of gold mining activities are stronger drivers (77 % model support) of Hg accumulation than environmental exposure (23 %) constitutes a novel contribution to a growing body of literature on Hg contamination. Our findings provide additional evidence of high Hg exposure risks to neotropical human and top carnivore populations subject to the impacts of ASGM, which depend on freshwater ecosystems undergoing gradual degradation. The documented spatial variation in Hg accumulation and increased Hg levels in carnivorous fish should serve as a warning to human communities in Madre de Dios to avoid the proximity of high-intensity gold mining areas and minimize local carnivorous fish consumption.

Alcohol, drugs, and road traffic injuries in an emergency department in Mexico City.

There have been no studies reporting on the use of biological specimens in Mexico to analyze the prevalence of alcohol and drug use among Emergency Department (ED) patients with a road traffic injury (RTI). We report here on a sample of 304 adult patients, admitted to the ED of a public hospital in Mexico City from January to April 2022, after being involved in an RTI. Patients gave informed consent for a breath test measuring breath alcohol concentration (BAC) and a saliva screening test for six classes of drugs (amphetamines, barbiturates, benzodiazepines, cannabis, cocaine, and methamphetamine). We found that at least one in every four patients (27.6%) had traces of alcohol or drugs in their body upon arrival in the ED. The breath test found a positive BAC in 16.1% of the sample; the most common substances detected in saliva were amphetamines or methamphetamine (8.6%), followed by cocaine (7.0%) and cannabis (6.9%). Only a few variables differentiated those with positive BAC from those with negative BAC (male, arriving on a weekend day or night, and arriving by ambulance), and even fewer variables differentiated those testing positives for drugs than those testing negative (less than 13 years of education and drivers of cars, bicycles, or other vehicles). While alcohol continues to be the single most used substance, our findings indicate that stimulants are of great concern. Since those testing positives for alcohol or drugs are so similar in their demographic pattern to those testing negative, the introduction of biological testing as a routine practice in the ED is highly recommended. Routine testing makes it possible to provide the patient with the best treatment and is also the best way to assess substance use.

Are Enterobacteriaceae and Enterococcus Isolated from Powdered Infant Formula a Hazard for Infants? A Genomic Analysis.

Powdered infant formulas (PIF) are the most used dietary substitutes that are used in order to supplement breastfeeding. However, PIF are not sterile and can be contaminated with different microorganisms. The objective of this study was to genomically characterize Enterobacteriaceae (ENT) and Enterococcus strains that were isolated from PIF. Strains were identified by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) and whole-genome sequencing (WGS). Genomic typing, detection of virulence, and resistance profiles and genes were performed with the Ridom SeqSphere+ software; the comprehensive antibiotic resistance database (CARD) platform; ResFinder and PlasmidFinder tools; and by the disk diffusion method. Nineteen isolates from PIF were analyzed, including ENT such as Kosakonia cowanii, Enterobacter hormaechei, Franconibacter helveticus, Mixta calida, and lactic acid bacteria such as Enterococcus faecium. The strains exhibited resistance to beta-lactams, cephalosporins, and macrolides. Resistance genes such as AcrAB-TolC, marA, msbA, knpEF, oqxAB, fosA, blaACT-7, blaACT-14,qacJ, oqxAB,aac(6')-Ii, and msr(C); and virulence genes such as astA, cheB, cheR, ompA ompX, terC, ironA, acm, and efaAfm, adem were also detected. All the analyzed strains possessed genes that produced heat-shock proteins, such as IbpA and ClpL. In PIF, the presence of ENT and Enterococcus that are multiresistant to antibiotics-together with resistance and virulence genes-pose a health risk for infants consuming these food products.

Maternal Factors Associated with Non-Exclusive Breastfeeding in Haitian Immigrant Women in Southern Chile.

There is limited knowledge concerning factors that affect non-exclusive breastfeeding (NEBF) practices in immigrant populations, especially in Latin America. The objective of the present study was to determine the association between maternal factors and the prevalence of NEBF in Haitian immigrant women in southern Chile. This was an analytical cross-sectional study. The probabilistic sample consisted of 173 Haitian women who gave informed consent. Sociodemographic and dietary-nutritional information was collected from all participants. Bivariate (χ2) and multivariate (logistic regression) inferential statistics were applied. All analyses were performed with the STATA 16.0 statistical software, and the significance level was established as α < 0.05. The prevalence of EBF at 6 mo was 54.3%. Maternal factors associated with a lower prevalence of EBF were not having permanent residency (OR: 2.34, CI: 2.18−2.83), residency <12 months (OR: 2.23, CI: 2.09−2.78), limited knowledge of breastfeeding (OR: 1.96; CI: 1.81−2.27), and low educational attainment (OR: 1.78; CI: 1.61−2.11). The protective factors were employment (OR: 0.36, CI: 0.28−0.40), access to basic services (OR: 0.32; CI: 0.22−0.48), and Spanish proficiency (OR: 0.29; CI: 0.20−0.51). Haitian immigrant women without legal residency, recently arrived, with low educational attainment, and poor knowledge of breastfeeding have more risk of not providing exclusive breastfeeding. Targeted interventions for mothers with these risk factors may help improve EBF rates.

Malnutrición por exceso en niños de 5 a 10 años y su asociación con el estado nutricional pre y gestacional, lactancia materna y patología materna

La malnutrición por exceso es un problema de salud pública relevante, con origen multifactorial y factores asociados como el estado nutricional pregestacional (ENP), durante y post gestación, ganancia excesiva de peso (EGP) gestacional y duración de la lactancia materna. El objetivo del estudio fue evaluar la malnutrición por exceso en niños de 5 a 10 años y su asociación con el estado nutricional pre y gestacional, lactancia materna exclusiva (LME) y patologías maternas. Se realizó un estudio con 213 niños y niñas estudiantes de 5 a 10 años. Los datos biosociodemográficos se obtuvieron mediante entrevistas con las madres y datos maternos previos y el embarazo con el carné de control. El estimó estado nutricional se estimó con z-score del peso para la talla e IMC/E, según sexo. Para los análisis estadísticos se usó el software STATA 15.0. El 56% de los estudiantes tuvo malnutrición por exceso. Un 53,1% de las madres presentó malnutrición por exceso previo al embarazo que aumentó a 74,6% post gestación. El 71,4% de los niños tuvo una LME ≥6 meses. Se relacionó estadísticamente la edad materna ≥35 años (p= 0,044) y la malnutrición por exceso pregestacional (p= 0,014). La edad materna ≥35 años aumentó el riesgo de malnutrición por exceso casi dos veces (OR= 1,78; IC: 1,029-3,046), al igual que el ENP (OR= 2,11; IC: 1,193-3,693) y en patologías maternas (OR= 1,41; IC: 1, 073-2,694). En conclusión, los niños preescolares de 5 a 10 años con factores de edad materna ≥35, ENP y patologías maternas tuvieron entre 1,4 y 2,11 veces más riesgo de presentar malnutrición por exceso comparado con aquellos niños sin estos factores.

Overnutrition is a relevant public health problem with a multifactorial origin. Associated factors include maternal nutritional status before, during, and after gestation, excessive gestational weight gain, and breastfeeding duration. The objective of the study was to evaluate the association between overnutrition in children aged 5-10 years and pre- and gestational nutritional status, exclusive breastfeeding, and maternal pathologies. The study consisted of 213 schoolchildren aged 5-10 years. Bio-socio-demographic data were obtained through interviews with mothers, previous maternal data and pregnancy follow-up records. Nutritional status was measured using weight-for-height and BMI z-score by age and sex. The STATA 15.0 software was used for statistical analysis. A total of 56% of the schoolchildren had overnutrition. Meanwhile, 53.1% of the mothers demonstrated pre-pregnancy overnutrition, which increased to 74.6% post-pregnancy. It was found that 71.4% of schoolchildren had experienced exclusive breastfeeding ≥6 months. Maternal age ≥35 years (p= 0.044) and pre-gestational nutritional status (p= 0.014) were statistically related. Maternal age ≥35 years increased overnutrition by almost two-fold (OR= 1.78; IC: 1.029-3.046), as did pre-gestational nutritional status (OR= 2.11; IC: 1.193-3.693) and maternal pathologies (OR= 1.41; IC: 1.073-2.694). In conclusion, schoolchildren aged 5-10 years with mothers ≥35 years of age, who had overnutrition in the pre-gestational period, and a pathology were 1.4 to 2.44 times more at risk of developing overnutrition than children without these factors.

Association between Maternal Factors, Preterm Birth, and Low Birth Weight of Chilean Singletons.

There has been an increase in preterm (PT) births in Western countries in recent years, which is associated with low-birthweight (LBW) children. The aim of this study was to determine the association between maternal factors and PT and LBW Chilean newborns. Methods: This was an analytical cross-sectional study of a national sample of 903,847 newborns and their mothers. The newborn gestational age, birth weight, maternal age, marital status, education, employment situation, and residence were analyzed. A multivariate logistic regression model was applied (α = 0.05) (STATA v.15). The prevalence was 6.8% and 5.0% for PT and LBW, respectively. The probability of the newborns being PT and LBW was 1.18 and 1.22 times if their mothers had <12 years of education and 1.38 and 1.29 times if the mothers were ≥35 years old, respectively. Mothers with <12 years education and ≥35 years were risk factors for PT and LBW newborns. Maternal educational attainment was a protective factor for the Chilean newborns, and a maternal age ≥35 years was a risk factor for PT and LBW.

Genomic Characterization of Cronobacter spp. and Salmonella spp. Strains Isolated From Powdered Infant Formula in Chile.

This study characterized five Cronobacter spp. and six Salmonella spp. strains that had been isolated from 155 samples of powdered infant formula (PIF) sold in Chile and manufactured in Chile and Mexico in 2018-2020. Two strains of Cronobacter sakazakii sequence type (ST) ST1 and ST31 (serotypes O:1 and O:2) and one strain of Cronobacter malonaticus ST60 (O:1) were identified. All Salmonella strains were identified as Salmonella Typhimurium ST19 (serotype O:4) by average nucleotide identity, ribosomal multilocus sequence typing (rMLST), and core genome MLST (cgMLST). The C. sakazakii and C. malonaticus isolates were resistant to cephalothin, whereas the Salmonella isolates were resistant to oxacillin and ampicillin. Nineteen antibiotic resistance genes were detected in the C. sakazakii and C. malonaticus isolates; the most prevalent were mcr-9.1, blaCSA , and blaCMA . In Salmonella, 30 genes encoding for aminoglycoside and cephalosporin resistance were identified, including aac(6')-Iaa, ß-lactamases ampH, ampC1, and marA. In the Cronobacter isolates, 32 virulence-associated genes were detected by WGS and clustered as flagellar proteins, outer membrane proteins, chemotaxis, hemolysins, invasion, plasminogen activator, colonization, transcriptional regulator, survival in macrophages, use of sialic acid, and toxin-antitoxin genes. In the Salmonella strains, 120 virulence associated genes were detected, adherence, magnesium uptake, resistance to antimicrobial peptides, secretion system, stress protein, toxin, resistance to complement killing, and eight pathogenicity islands. The C. sakazakii and C. malonaticus strains harbored I-E and I-F CRISPR-Cas systems and carried Col(pHHAD28) and IncFIB(pCTU1) plasmids, respectively. The Salmonella strains harbored type I-E CRISPR-Cas systems and carried IncFII(S) plasmids. The presence of C. sakazakii and Salmonella in PIF is a health risk for infants aged less than 6 months. For this reason, sanitary practices should be reinforced for its production and retail surveillance.

Obese Women Have a High Carbohydrate Intake without Changes in the Resting Metabolic Rate in the Luteal Phase.

Hormonal changes are caused by the menstrual cycle phases, which influence resting metabolic rate and eating behavior. The aim of the study was to determine resting metabolic rate (RMR) and its association with dietary intake according to the menstrual cycle phase in lean and obese Chilean women. This cross-sectional analytical study included 30 adult women (15 lean and 15 with obesity). Body composition was measured with a tetrapolar bioelectrical impedance meter. Nutritional status was determined by adiposity. A 24-h recall of three nonconsecutive days verifies dietary intake. The RMR was measured by indirect calorimetry. All measurements were performed in both the follicular and luteal phases of the menstrual cycle. Statistical analyses were performed with STATA software at a significance level, which was α = 0.05. The RMR (ß = 121.6 kcal/d), temperature (ß = 0.36 °C), calorie intake (ß = 317.1 kcal/d), and intake of lipids (ß = 13.8 g/d) were associated with the luteal phase in lean women. Only extracellular water (ß = 1.11%) and carbohydrate consumption (ß = 45.2 g/d) were associated in women with obesity. Lean women showed increased RMR, caloric intake, and lipid intake during the luteal phase. For women with obesity, carbohydrate intake increased but not RMR.

Association of Prematurity and Low Birth Weight with Gestational Exposure to PM2.5 and PM10 Particulate Matter in Chileans Newborns.

Fetal growth can be affected by gestational exposure to air pollution. The aim of the study was to determine the association between prematurity and low birth weight (LBW) with gestational exposure to PM2.5 and PM10 particulate matter in Chileans newborns. This cross-sectional analytical study included 595,369 newborns. Data were extracted from the live newborn records of the Chilean Ministry of Health. Sex, gestational age, birth weight, and living variables were analyzed. We used the Air Quality Information System of the Chilean Ministry of the Environment to obtain mean PM2.5 and PM10 emissions. A multivariate logistic regression model was performed with STATA 15.0 software at α < 0.05. Prevalence was 7.4% prematurity and 5.5% LBW. Mean PM2.5 and PM10 concentrations were 25.5 µg/m3 and 55.3 µg/m3, respectively. PM2.5 was associated with an increased the risk of LBW (OR: 1.031; 95%CI: 1.004−1.059) when exposure occurred in the second trimester, while PM10 affected the whole pregnancy. In addition, PM10 exposure in any gestational trimester was associated with an increased the risk of prematurity. The PM10 particulate matter was associated with both prematurity and LBW in all of the trimesters of exposure. The PM2.5 particulate matter was only associated with LBW when exposure occurred in the second gestational trimester.

Tet1 Suppresses p21 to Ensure Proper Cell Cycle Progression in Embryonic Stem Cells.

Ten eleven translocation 1 (Tet1) is a DNA dioxygenase that promotes DNA demethylation by oxidizing 5-methylcytosine. It can also partner with chromatin-activating and repressive complexes to regulate gene expressions independent of its enzymatic activity. Tet1 is highly expressed in embryonic stem cells (ESCs) and regulates pluripotency and differentiation. However, its roles in ESC cell cycle progression and proliferation have not been investigated. Using a series of Tet1 catalytic mutant (Tet1m/m), knockout (Tet1-/-) and wild type (Tet1+/+) mouse ESCs (mESCs), we identified a non-catalytic role of Tet1 in the proper cell cycle progression and proliferation of mESCs. Tet1-/-, but not Tet1m/m, mESCs exhibited a significant reduction in proliferation and delayed progression through G1. We found that the cyclin-dependent kinase inhibitor p21/Cdkn1a was uniquely upregulated in Tet1-/- mESCs and its knockdown corrected the slow proliferation and delayed G1 progression. Mechanistically, we found that p21 was a direct target of Tet1. Tet1 occupancy at the p21 promoter overlapped with the repressive histone mark H3K27me3 as well as with the H3K27 trimethyl transferase PRC2 component Ezh2. A loss of Tet1, but not loss of its catalytic activity, significantly reduced the enrichment of Ezh2 and H3K27 trimethylation at the p21 promoter without affecting the DNA methylation levels. We also found that the proliferation defects of Tet1-/- mESCs were independent of their differentiation defects. Together, these findings established a non-catalytic role for Tet1 in suppressing p21 in mESCs to ensure a rapid G1-to-S progression, which is a key hallmark of ESC proliferation. It also established Tet1 as an epigenetic regulator of ESC proliferation in addition to its previously defined roles in ESC pluripotency and differentiation.

Idax and Rinf facilitate expression of Tet enzymes to promote neural and suppress trophectodermal programs during differentiation of embryonic stem cells.

The Inhibitor of disheveled and axin (Idax) and its ortholog the Retinoid inducible nuclear factor (Rinf) are DNA binding proteins with nuclear and cytoplasmic functions. Rinf is expressed in embryonic stem cells (ESCs) where it regulates transcription of the Ten-eleven translocation (Tet) enzymes, promoting neural and suppressing mesendoderm/trophectoderm differentiation. Here, we find that Idax, which is not expressed in ESCs, is induced upon differentiation. Like Rinf, Idax facilitates neural and silences trophectodermal programs. Individual or combined loss of Idax and Rinf led to downregulation of neural and upregulation of trophectoderm markers during differentiation of ESCs to embryoid bodies as well as during directed differentiation of ESCs to neural progenitor cells (NPCs) and trophoblast-like cells. These defects resemble those of Tet-deficient ESCs. Consistently, Tet genes are direct targets of Idax and Rinf, and loss of Idax and Rinf led to downregulation of Tet enzymes during ESC differentiation to NPCs and trophoblast-like cells. While Idax and Rinf single and double knockout (DKO) mice were viable and overtly normal, DKO embryos had reduced expression of several NPC markers in embryonic forebrains and deregulated expression of selected trophoblast markers in placentas. NPCs derived from DKO forebrains had reduced self-renewal while DKO placentas had increased junctional zone and reduced labyrinth layers. Together, our findings establish Idax and Rinf as regulators of Tet enzymes for proper differentiation of ESCs.

Definitive hematopoietic stem cells minimally contribute to embryonic hematopoiesis.

Hematopoietic stem cells (HSCs) are rare cells that arise in the embryo and sustain adult hematopoiesis. Although the functional potential of nascent HSCs is detectable by transplantation, their native contribution during development is unknown, in part due to the overlapping genesis and marker gene expression with other embryonic blood progenitors. Using single-cell transcriptomics, we define gene signatures that distinguish nascent HSCs from embryonic blood progenitors. Applying a lineage-tracing approach to selectively track HSC output in situ, we find significantly delayed lymphomyeloid contribution. An inducible HSC injury model demonstrates a negligible impact on larval lymphomyelopoiesis following HSC depletion. HSCs are not merely dormant at this developmental stage, as they showed robust regeneration after injury. Combined, our findings illuminate that nascent HSCs self-renew but display differentiation latency, while HSC-independent embryonic progenitors sustain developmental hematopoiesis. Understanding these differences could improve de novo generation and expansion of functional HSCs.

Draft Genome Sequences of Seven Cronobacter sakazakii Strains Carrying the mcr 9.1 Gene Isolated in Chile.

Cronobacter sakazakii is a pathogen that causes severe diseases such as meningitis and necrotizing enterocolitis in infants under 12 months, associated with the consumption of contaminated rehydrated powdered infant formula (PIF). We present seven C. sakazakii genome sequences isolated from PIF and dairy products in Chile in 2017.