Gait Characteristics and Cognitive Function in Middle-Aged Adults with and without Type 2 Diabetes Mellitus: Data from ENBIND.

Type 2 Diabetes Mellitus (T2DM) in midlife is associated with a greater risk of dementia in later life. Both gait speed and spatiotemporal gait characteristics have been associated with later cognitive decline in community-dwelling older adults. Thus, the assessment of gait characteristics in uncomplicated midlife T2DM may be important in selecting-out those with T2DM at greatest risk of later cognitive decline. We assessed the relationship between Inertial Motion Unit (IMUs)-derived gait characteristics and cognitive function assessed via Montreal Cognitive Assessment (MoCA)/detailed neuropsychological assessment battery (CANTAB) in middle-aged adults with and without uncomplicated T2DM using both multivariate linear regression and a neural network approach. Gait was assessed under (i) normal walking, (ii) fast (maximal) walking and (iii) cognitive dual-task walking (reciting alternate letters of the alphabet) conditions. Overall, 138 individuals were recruited (n = 94 with T2DM; 53% female, 52.8 ± 8.3 years; n = 44 healthy controls, 43% female, 51.9 ± 8.1 years). Midlife T2DM was associated with significantly slower gait velocity on both slow and fast walks (both p < 0.01) in addition to a longer stride time and greater gait complexity during normal walk (both p < 0.05). Findings persisted following covariate adjustment. In analyzing cognitive performance, the strongest association was observed between gait velocity and global cognitive function (MoCA). Significant associations were also observed between immediate/delayed memory performance and gait velocity. Analysis using a neural network approach did not outperform multivariate linear regression in predicting cognitive function (MoCA) from gait velocity. Our study demonstrates the impact of uncomplicated T2DM on gait speed and gait characteristics in midlife, in addition to the striking relationship between gait characteristics and global cognitive function/memory performance in midlife. Further studies are needed to evaluate the longitudinal relationship between midlife gait characteristics and later cognitive decline, which may aid in selecting-out those with T2DM at greatest-risk for preventative interventions.

Copper Tolerance and Characterization of a Copper-Responsive Operon, copYAZ, in an M1T1 Clinical Strain of Streptococcus pyogenes.

UNLABELLED: Infection with Streptococcus pyogenes is associated with a breadth of clinical manifestations ranging from mild pharyngitis to severe necrotizing fasciitis. Elevated levels of intracellular copper are highly toxic to this bacterium, and thus, the microbe must tightly regulate the level of this metal ion by one or more mechanisms, which have, to date, not been clearly defined. In this study, we have identified two virulence mechanisms by which S. pyogenes protects itself against copper toxicity. We defined a set of putative genes, copY (for a regulator), copA (for a P1-type ATPase), and copZ (for a copper chaperone), whose expression is regulated by copper. Our results indicate that these genes are highly conserved among a range of clinical S. pyogenes isolates. The copY, copA, and copZ genes are induced by copper and are transcribed as a single unit. Heterologous expression assays revealed that S. pyogenes CopA can confer copper tolerance in a copper-sensitive Escherichia coli mutant by preventing the accumulation of toxic levels of copper, a finding that is consistent with a role for CopA in copper export. Evaluation of the effect of copper stress on S. pyogenes in a planktonic or biofilm state revealed that biofilms may aid in protection during initial exposure to copper. However, copper stress appears to prevent the shift from the planktonic to the biofilm state. Therefore, our results indicate that S. pyogenes may use several virulence mechanisms, including altered gene expression and a transition to and from planktonic and biofilm states, to promote survival during copper stress. IMPORTANCE: Bacterial pathogens encounter multiple stressors at the host-pathogen interface. This study evaluates a virulence mechanism(s) utilized by S. pyogenes to combat copper at sites of infection. A better understanding of pathogen tolerance to stressors such as copper is necessary to determine how host-pathogen interactions impact bacterial survival during infections. These insights may lead to the identification of novel therapeutic targets that can be used to address antibiotic resistance.

Eradicating group A streptococcus bacteria and biofilms using functionalised multi-wall carbon nanotubes.

PURPOSE: The aim of this study was to demonstrate that multi-wall carbon nanotubes can be functionalised with antibodies to group A streptoccocus (GAS) for targeted photothermal ablation of planktonic and biofilm residing bacteria. MATERIALS AND METHODS: Antibodies for GAS were covalently attached to carboxylated multi-wall carbon nanotubes and incubated with either planktonic or biofilm GAS. Bacterium was then exposed to 1.3 W/cm(2) of 800 nm light for 10-120 s, and then serially diluted onto agar plates from which the number of colony forming units was determined. Photothermal ablation of GAS on the surface of full thickness ex vivo porcine skin and histological sectioning were done to examine damage in adjacent tissue. RESULTS: Approximately 14% of the GAS antibody-functionalised nanotubes attached to the bacterium, and this amount was found to be capable of inducing photothermal ablation of GAS upon exposure to 1.3 W/cm(2) of 800 nm light. Cell viability was not decreased upon exposure to nanotubes or infrared light alone. Compared to carboxylated multi-wall carbon nanotubes, antibody-labelled nanotubes enhanced killing in both planktonic and biofilm GAS in conjunction with infrared light. Analysis of GAS photothermally ablated in direct contact with ex vivo porcine skin shows that heat sufficient for killing GAS remains localised and does not cause collateral damage in tissue adjacent to the treated area. CONCLUSIONS: The results of this study support the premise that carbon nanotubes may be effectively utilised as highly localised photothermal agents with the potential for translation into the clinical treatment of bacterial infections of soft tissue.

Geographic variations of pre-exposure prophylaxis reversal and abandonment among United States counties.

OBJECTIVE: In the United States, one in five newly insurer-approved pre-exposure prophylaxis (PrEP) prescriptions are reversed with over 70% of those reversed, being abandoned. Given the Ending the HIV Epidemic (EHE) initiative's goals, we assessed geographic variations of PrEP reversal and abandonment across EHE and non-EHE counties in the United States. DESIGN: This was a cross-sectional analysis of secondary data. METHODS: Data were collected from Symphony Analytics for adults 18 years and older, with a newly prescribed PrEP claim. Using the proportion of PrEP prescriptions by county, hotspot analysis was conducted utilizing Getis Ord Gi∗ statistics stratified by EHE and non EHE counties. Multivariable logistic regression was used to identify factors associated with residing in hotspots of PrEP reversal or PrEP abandonments. RESULTS: Across 516 counties representing 36,204 patients, the overall PrEP reversal rate was 19.4%, whereas the PrEP abandonment rate was 13.7%. Reversals and abandonments were higher for non-EHE (22.7 and 17.1%) than EHE (15.6 and 10.5%) counties. In both EHE and non-EHE counties, younger age, less education, females, and an out-of-pocket cost of greater than $100, were significantly associated with greater likelihood of residing in hotspots of PrEP reversal or abandonment, while Hispanics, Medicaid recipients, and an out-of-pocket cost of $10 or less had lower likelihood of residing in hotspots of reversal and abandonment. CONCLUSION: Findings indicate the need for implementation of focused interventions to address disparities observed in PrEP reversal and abandonment. Moreover, to improve primary PrEP adherence, national PrEP access programs should streamline and improve PrEP accessibility across different geographic jurisdictions.

Geographic mobility and its impact on sexual health and ongoing HIV transmission among migrant latinx men who have sex with men.

An understudied social process that may determine variable HIV risk, testing, and linkage to care is geographic mobility, including immigration as well as short-term mobility, especially among sexual minority populations. We aimed to assess how geographic mobility over the lifecourse between Latin America and the U.S., and within the U.S., was linked to sexual risk and health behaviors among Latinx migrant men who have sex with men (MSM) in San Bernardino County, California. Qualitative analysis of 16 semi-structured interviews revealed four major domains of influence on participants' sexual risk behaviors. At the micro level, these included social environment/interpersonal factors (e.g., family and peer support) and geographic factors and pathways (e.g., migration journey to the U.S.). At the macro level data centered on cultural factors (e.g., gender norms in home country) and structural factors (e.g., HIV healthcare). Our results can illuminate and promote effective health policies and HIV reduction efforts for Latinx migrant MSM in metro areas.

Novel Location-Based Survey Using Cognitive Interviews to Assess Geographic Networks and Hotspots of Sex and Drug Use: Implementation and Validation Study.

BACKGROUND: The Ending the HIV Epidemic initiative in the United States relies on HIV hotspots to identify where to geographically target new resources, expertise, and technology. However, interventions targeted at places with high HIV transmission and infection risk, not just places with high HIV incidence, may be more effective at reducing HIV incidence and achieving health equity. OBJECTIVE: We described the implementation and validation of a web-based activity space survey on HIV risk behaviors. The survey was intended to collect geographic information that will be used to map risk behavior hotspots as well as the geography of sexual networks in Los Angeles County. METHODS: The survey design team developed a series of geospatial questions that follow a 3-level structure that becomes more geographically precise as participants move through the levels. The survey was validated through 9 cognitive interviews and iteratively updated based on participant feedback until the saturation of topics and technical issues was reached. RESULTS: In total, 4 themes were identified through the cognitive interviews: functionality of geospatial questions, representation and accessibility, privacy, and length and understanding of the survey. The ease of use for the geospatial questions was critical as many participants were not familiar with mapping software. The inclusion of well-known places, landmarks, and road networks was critical for ease of use. The addition of a Google Maps interface, which was familiar to many participants, aided in collecting accurate and precise location information. The geospatial questions increased the length of the survey and warranted the inclusion of features to simplify it and speed it up. Using nicknames to refer to previously entered geographic locations limited the number of geospatial questions that appeared in the survey and reduced the time taken to complete it. The long-standing relationship between participants and the research team improved comfort to disclose sensitive geographic information related to drug use and sex. Participants in the cognitive interviews highlighted how trust and inclusive and validating language in the survey alleviated concerns related to privacy and representation. CONCLUSIONS: This study provides promising results regarding the feasibility of using a web-based mapping survey to collect sensitive location information relevant to ending the HIV epidemic. Data collection at several geographic levels will allow for insights into spatial recall of behaviors as well as future sensitivity analysis of the spatial scale of hotspots and network characteristics. This design also promotes the privacy and comfort of participants who provide location information for sensitive topics. Key considerations for implementing this type of survey include trust from participants, community partners, or research teams to overcome concerns related to privacy and comfort. The implementation of similar surveys should consider local characteristics and knowledge when crafting the geospatial components.

Is alcohol-based hand disinfection equivalent to surgical scrub before placing a central venous catheter?

BACKGROUND: Waterless antiseptic surgical hand scrub (1% chlorhexidine gluconate and 61% ethyl alcohol, Avagard™; 3M Health Care, St. Paul, MN), alcohol-only cleanser (62% ethyl alcohol), and traditional surgical scrub (5-minute scrub with 4% chlorhexidine soap using a sterile scrub brush with water) are techniques used for hand cleansing and disinfection. We hypothesized that alcohol-only cleanser and waterless antiseptic scrub (Avagard) would be as effective as a traditional surgical scrub for hand cleansing before placement of central venous catheters. METHODS: Fingers of subjects were plate-cultured for 24 hours after 5 methods of hand cleansing: method 1: traditional surgical scrub (n = 49 plates produced by 14 subjects); method 2: traditional surgical scrub (5-minute scrub with water, brush, and 4% chlorhexidine soap) followed by a 15-minute break, then alcohol-only cleanser (62% alcohol) (n = 49 plates produced by 14 subjects); method 3: alcohol-only cleanser alone (n = 49 plates produced by 14 subjects); method 4: alcohol-only cleanser (62% alcohol), followed by a 15-minute break, then traditional surgical scrub (5-minute scrub with brush, and 4% chlorhexidine soap with water) (n = 49 plates produced by 14 subjects); and method 5: waterless surgical scrub (Avagard) alone (n = 116 plates produced by 38 subjects). The 15-minute break was introduced to allow a short period of recontamination, and to test for residual effects from prior cleansing. RESULTS: Alcohol-only cleanser alone (method 3) was significantly less effective than the traditional surgical scrub (method 1) (P < 0.001; 82% plate growth). Waterless surgical scrub (Avagard) (method 5) had a 0% observed difference (95% confidence interval [CI]: -14% to 11%) compared with the traditional 5-minute scrub (method 1) (P = 0.99; 16% plate growth). When a traditional surgical scrub was used first followed by a 15-minute period of recontamination, there was a 6% observed difference in method 2 from reference (method 1) (95% CI: -10% to 22%), and 0% observed difference in method 4 from reference (95% CI: -15% to 15%). CONCLUSION: As the initial cleansing method, the alcohol-only cleanser (method 3) was significantly less effective than the traditional surgical scrub (method 1) (P < 0.001).

Detection of group A Streptococcus in tonsils from pediatric patients reveals high rate of asymptomatic streptococcal carriage.

BACKGROUND: Group A Streptococcus (GAS) causes acute tonsillopharyngitis in children, and approximately 20% of this population are chronic carriers of GAS. Antibacterial therapy has previously been shown to be insufficient at clearing GAS carriage. Bacterial biofilms are a surface-attached bacterial community that is encased in a matrix of extracellular polymeric substances. Biofilms have been shown to provide a protective niche against the immune response and antibiotic treatments, and are often associated with recurrent or chronic bacterial infections. The objective of this study was to test the hypothesis that GAS is present within tonsil tissue at the time of tonsillectomy. METHODS: Blinded immunofluorescent and histological methods were employed to evaluate palatine tonsils from children undergoing routine tonsillectomy for adenotonsillar hypertrophy or recurrent GAS tonsillopharyngitis. RESULTS: Immunofluorescence analysis using anti-GAS antibody was positive in 11/30 (37%) children who had tonsillectomy for adenotonsillar hypertrophy and in 10/30 (33%) children who had tonsillectomy for recurrent GAS pharyngitis. Fluorescent microscopy with anti-GAS and anti-cytokeratin 8 & 18 antibodies revealed GAS was localized to the tonsillar reticulated crypts. Scanning electron microscopy identified 3-dimensional communities of cocci similar in size and morphology to GAS. The characteristics of these communities are similar to GAS biofilms from in vivo animal models. CONCLUSION: Our study revealed the presence of GAS within the tonsillar reticulated crypts of approximately one-third of children who underwent tonsillectomy for either adenotonsillar hypertrophy or recurrent GAS tonsillopharyngitis at the Wake Forest School of Medicine. TRIAL REGISTRATION: The tissue collected was normally discarded tissue and no patient identifiers were collected. Thus, no subjects were formally enrolled.

One third of middle ear effusions from children undergoing tympanostomy tube placement had multiple bacterial pathogens.

BACKGROUND: Because previous studies have indicated that otitis media may be a polymicrobial disease, we prospectively analyzed middle ear effusions of children undergoing tympanostomy tube placement with multiplex polymerase chain reaction for four otopathogens. METHODS: Middle ear effusions from 207 children undergoing routine tympanostomy tube placement were collected and were classified by the surgeon as acute otitis media (AOM) for purulent effusions and as otitis media with effusion (OME) for non-purulent effusions. DNA was isolated from these samples and analyzed with multiplex polymerase chain reaction for Haemophilus influenzae, Streptococcus pneumoniae, Alloiococcus otitidis, and Moraxella catarrhalis. RESULTS: 119 (57%) of 207 patients were PCR positive for at least one of these four organisms. 36 (30%) of the positive samples indicated the presence of more than one bacterial species. Patient samples were further separated into 2 groups based on clinical presentation at the time of surgery. Samples were categorized as acute otitis media (AOM) if pus was observed behind the tympanic membrane. If no pus was present, samples were categorized as otitis media with effusion (OME). Bacteria were identified in most of the children with AOM (87%) and half the children with OME (51%, p < 0.001). A single bacterial organism was detected in middle ear effusions from children with AOM more often than those with OME (74% versus 33%, p < 0.001). Haemophilus influenzae was the predominant single organism and caused 58% of all AOM in this study. Alloiococcus otitidis and Moraxella catarrhalis were more frequently identified in middle ear effusions than Streptococcus pneumoniae. CONCLUSIONS: Haemophilus influenzae, Streptococcus pneumoniae, Alloiococcus otitidis, and Moraxella catarrhalis were identified in the middle ear effusions of some patients with otitis media. Overall, we found AOM is predominantly a single organism infection and most commonly from Haemophilus influenzae. In contrast, OME infections had a more equal distribution of single organisms, polymicrobial entities, and non-bacterial agents.

The streptococcal collagen-like protein-1 (Scl1) is a significant determinant for biofilm formation by group A Streptococcus.

BACKGROUND: Group A Streptococcus (GAS) is a human-specific pathogen responsible for a number of diseases characterized by a wide range of clinical manifestations. During host colonization GAS-cell aggregates or microcolonies are observed in tissues. GAS biofilm, which is an in vitro equivalent of tissue microcolony, has only recently been studied and little is known about the specific surface determinants that aid biofilm formation. In this study, we demonstrate that surface-associated streptococcal collagen-like protein-1 (Scl1) plays an important role in GAS biofilm formation. RESULTS: Biofilm formation by M1-, M3-, M28-, and M41-type GAS strains, representing an intraspecies breadth, were analyzed spectrophotometrically following crystal violet staining, and characterized using confocal and field emission scanning electron microscopy. The M41-type strain formed the most robust biofilm under static conditions, followed by M28- and M1-type strains, while the M3-type strains analyzed here did not form biofilm under the same experimental conditions. Differences in architecture and cell-surface morphology were observed in biofilms formed by the M1- and M41-wild-type strains, accompanied by varying amounts of deposited extracellular matrix and differences in cell-to-cell junctions within each biofilm. Importantly, all Scl1-negative mutants examined showed significantly decreased ability to form biofilm in vitro. Furthermore, the Scl1 protein expressed on the surface of a heterologous host, Lactococcus lactis, was sufficient to induce biofilm formation by this organism. CONCLUSIONS: Overall, this work (i) identifies variations in biofilm formation capacity among pathogenically different GAS strains, (ii) identifies GAS surface properties that may aid in biofilm stability and, (iii) establishes that the Scl1 surface protein is an important determinant of GAS biofilm, which is sufficient to enable biofilm formation in the heterologous host Lactococcus. In summary, the GAS surface adhesin Scl1 may have an important role in biofilm-associated pathogenicity.

Characterization of the N-acetyl-α-D-glucosaminyl l-malate synthase and deacetylase functions for bacillithiol biosynthesis in Bacillus anthracis .

Bacillithiol (Cys-GlcN-malate, BSH) has recently been identified as a novel low-molecular weight thiol in Bacillus anthracis, Staphylococcus aureus, and several other Gram-positive bacteria lacking glutathione and mycothiol. We have now characterized the first two enzymes for the BSH biosynthetic pathway in B. anthracis, which combine to produce α-d-glucosaminyl l-malate (GlcN-malate) from UDP-GlcNAc and l-malate. The structure of the GlcNAc-malate intermediate has been determined, as have the kinetic parameters for the BaBshA glycosyltransferase (→GlcNAc-malate) and the BaBshB deacetylase (→GlcN-malate). BSH is one of only two natural products reported to contain a malyl glycoside, and the crystal structure of the BaBshA-UDP-malate ternary complex, determined in this work at 3.3 Å resolution, identifies several active-site interactions important for the specific recognition of l-malate, but not other α-hydroxy acids, as the acceptor substrate. In sharp contrast to the structures reported for the GlcNAc-1-d-myo-inositol-3-phosphate synthase (MshA) apo and ternary complex forms, there is no major conformational change observed in the structures of the corresponding BaBshA forms. A mutant strain of B. anthracis deficient in the BshA glycosyltransferase fails to produce BSH, as predicted. This B. anthracis bshA locus (BA1558) has been identified in a transposon-site hybridization study as required for growth, sporulation, or germination [Day, W. A., Jr., Rasmussen, S. L., Carpenter, B. M., Peterson, S. N., and Friedlander, A. M. (2007) J. Bacteriol. 189, 3296-3301], suggesting that the biosynthesis of BSH could represent a target for the development of novel antimicrobials with broad-spectrum activity against Gram-positive pathogens like B. anthracis. The metabolites that function in thiol redox buffering and homeostasis in Bacillus are not well understood, and we present a composite picture based on this and other recent work.

Loss of the group A Streptococcus regulator Srv decreases biofilm formation in vivo in an otitis media model of infection.

Group A Streptococcus (GAS) is a common causative agent of pharyngitis, but the role of GAS in otitis media is underappreciated. In this study, we sought to test the hypothesis that GAS colonizes the middle ear and establishes itself in localized, three-dimensional communities representative of biofilms. To test this hypothesis, the middle ears of chinchillas were infected with either a strain of GAS capable of forming biofilms in vitro (MGAS5005) or a strain deficient in biofilm formation due to the lack of the transcriptional regulator Srv (MGAS5005 Δsrv). Infection resulted in the formation of large, macroscopic structures within the middle ears of MGAS5005- and MGAS5005 Δsrv-infected animals. Plate counts, scanning electron microscopy, LIVE/DEAD staining, and Gram staining revealed a difference in the distributions of MGAS5005 versus MGAS5005 Δsrv in the infected samples. High numbers of CFU of MGAS5005 Δsrv were isolated from the middle ear effusion, and MGAS5005 Δsrv was found randomly distributed throughout the excised macroscopic structure. In contrast, MGAS5005 was found in densely packed microcolonies indicative of biofilms within the excised material from the middle ear. CFU levels of MGAS5005 from the effusion were significantly lower than that of MGAS5005 Δsrv early during the course of infection. Allelic replacement of the chromosomally encoded streptococcal cysteine protease (speB) in the MGAS5005 Δsrv background restored biofilm formation in vivo. Interestingly, our results suggest that GAS naturally forms a biofilm during otitis media but that biofilm formation is not required to establish infection following transbullar inoculation of chinchillas.

Lower Limb Graduated Compression Garments Modulate Autonomic Nervous System and Improve Post-Training Recovery Measured via Heart Rate Variability.

Prior studies have examined the benefits of graduated compression garments (GCG) with regards to diverse exercise regimens; however, the relationship between GCG and the autonomic nervous system (ANS) has not been fully explored. The aim of this study was to examine Heart Rate Variability (HRV) trends-a proxy for ANS modulation-in response to donning GCG during a progressive overload training regimen designed to induce overtraining. Ten college-aged male novice runners were recruited for the 8-week crossover study. After three weeks of monitored free living, participants were randomized and blinded to an intervention group that donned a lower-body GCG during a two-week exercise regimen or a control group that donned a visually identical but non-compressive sham during identical training. No significant difference in HRV was calculated by the natural logarithm of the root mean square of successive RR-interval differences (lnRMSSD) between the 3-week free-living baseline and GCG intervention periods (P = 0.3040). The mean lnRMSSD was greater during the free-living phase and GCG intervention compared to the sham placebo (P < 0.001 and <0.001 respectively). With regard to the daily fluctuation of lnRMSSD, no significant differences were found between free-living and intervention (P = 1.000). Conversely, the intervention period demonstrated reduced daily fluctuation of lnRMSSD relative to the Sham placebo group (P = 0.010). These novel findings posit that post training use of a commercially available graduated compression garment in novice runners may be effective in counteracting some deleterious effects from overtraining while attenuating its effects on vagally-mediated HRV.

Intensive glycemic control in gestational diabetes mellitus: a randomized controlled clinical feasibility trial.

BACKGROUND: Overweight and obese women with gestational diabetes mellitus are at increased risk for adverse perinatal outcomes, and they are also more likely to have suboptimal glycemic control. However, there is a paucity of data evaluating whether lower glycemic targets could improve outcomes. OBJECTIVE: To evaluate the feasibility of intensive glycemic control in overweight and obese women with gestational diabetes mellitus. MATERIALS AND METHODS: We randomized 60 overweight or obese women with gestational diabetes mellitus, diagnosed between 12 and 32 weeks' gestation to either intensive (fasting <90 mg/dL, 1 hour postprandial <120 mg/dL) or standard (fasting <95 mg/dL, 1 ho postprandial <140 mg/dL) glycemic targets. Maternal glucose was assessed in 2 ways: blinded continuous glucose monitors, worn for 5 days at 2 time points (at 12-32 weeks and again at 32-36 weeks), and self-monitored glucose measurement 4 times per day. All women underwent standardized dietary counseling, and medical therapy was prescribed as needed to achieve glycemic control. RESULTS: Between December 2015 and December 2017, we randomized 60 women to either intensive (n = 30) or standard (n = 30) glycemic control. Baseline characteristics including maternal age, body mass index, and gestational age at diagnosis were similar between the intensive and standard groups. Medical therapy was more common in women in the intensive group than those in the standard group (83 vs 57%, P = .02). Women in the intensive glycemic control group had lower glucose values as assessed by continuous glucose monitors at including 24-hour mean (-8.1; 95% confidence interval, -12.0 to -4.3 mg/dL; P < .0001) and 1-h postprandial (-11.8; 95% confidence interval, -19.7 to -3.9 mg/dL, P = .004) values. Hypoglycemia <60 mg/dL was uncommon and did not differ between groups. CONCLUSION: Intensive glycemic targets can be used in overweight and obese women with minimal hypoglycemia, and this approach results in improved glycemic control when compared to standard glycemic targets. Further studies are needed to determine whether intensive glycemic targets can improve maternal and neonatal outcomes in high-risk women with gestational diabetes mellitus. CLINICAL TRIAL IDENTIFIER: NCT02530866; clinicaltrials.gov.

The type III pantothenate kinase encoded by coaX is essential for growth of Bacillus anthracis.

In Bacillus anthracis, the novel type III pantothenate kinase (PanK(Ba); encoded by coaX) catalyzes the first committed step in coenzyme A biosynthesis. We have demonstrated by analyzing the growth characteristics of a conditional coaX mutant that PanK(Ba) is an essential enzyme, thus contributing to its validation as a new antimicrobial target.

Inactivation of the group A Streptococcus regulator srv results in chromosome wide reduction of transcript levels, and changes in extracellular levels of Sic and SpeB.

Group A Streptococcus is characterized by the ability to cause a diverse number of human infections including pharyngitis, necrotizing fasciitis, toxic shock syndrome, and acute rheumatic fever, yet the regulation of streptococcal genes involved in disease processes and survival in the host is not completely understood. Genome scale analysis has revealed a complex regulatory network including 13 two-component regulatory systems and more than 100 additional putative regulators, the majority of which remain uncharacterized. Among these is the streptococcal regulator of virulence, Srv, the first Group A Streptococcus member of the Crp/Fnr family of transcriptional regulators. Previous work demonstrated that the loss of srv resulted in a significant decrease in Group A Streptococcus virulence. To begin to define the gene products influenced by Srv, we combined microarray and two-dimensional gel electrophoresis analysis. Loss of srv results in a chromosome wide reduction of gene transcription and changes in the production of the extracellular virulence factors Sic (streptococcal inhibitor of complement) and SpeB (cysteine proteinase). Sic levels are reduced in the srv mutant, whereas the extracellular concentration and activity of SpeB is increased. These data link Srv to the increasingly complex GAS regulatory network.

Complex conditional control by pigeons in a continuous virtual environment.

We tested two pigeons in a continuously streaming digital environment. Using animation software that constantly presented a dynamic, three-dimensional (3D) environment, the animals were tested with a conditional object identification task. The correct object at a given time depended on the virtual context currently streaming in front of the pigeon. Pigeons were required to accurately peck correct target objects in the environment for food reward, while suppressing any pecks to intermixed distractor objects which delayed the next object's presentation. Experiment 1 established that the pigeons' discrimination of two objects could be controlled by the surface material of the digital terrain. Experiment 2 established that the pigeons' discrimination of four objects could be conjunctively controlled by both the surface material and topography of the streaming environment. These experiments indicate that pigeons can simultaneously process and use at least two context cues from a streaming environment to control their identification behavior of passing objects. These results add to the promise of testing interactive digital environments with animals to advance our understanding of cognition and behavior.

Macrolide-resistant Streptococcus pyogenes in the United States, 2002-2003.

BACKGROUND: Increased levels of macrolide-resistant Streptococcus pyogenes in focal regions of the United States have been reported. The purpose of this study was to determine the antimicrobial susceptibility of a large collection of S. pyogenes isolates from throughout the United States and to elucidate the mechanisms of resistance and genetic relatedness of macrolide-resistant isolates. METHODS: During 2002-2003, a total of 1885 S. pyogenes clinical isolates were obtained from 45 US medical centers. Susceptibility to penicillin, cefdinir, erythromycin, azithromycin, clarithromycin, clindamycin, telithromycin, and levofloxacin was determined. Macrolide resistance phenotypes were determined by double-disk diffusion, and macrolide resistance genotypes were determined by polymerase chain reaction and sequencing. All macrolide-resistant isolates and all isolates recovered from sterile sites were further characterized by pulsed-field gel electrophoresis (PFGE) and emm typing. RESULTS: The majority (85%) of isolates were pharyngeal. Resistance was detected to erythromycin (6.8% of isolates), azithromycin (6.9%), clarithromycin (6.6%), clindamycin (0.5%), telithromycin (0.2%), and levofloxacin (0.05%). The macrolide-resistance phenotype distribution was as follows: macrolide-lincosamide-streptogramin B (MLSB), 56% of isolates (inducible, 47%; constitutive, 9%); and M, 44%. The genotypes detected were as follows: ermA, 46% of isolates (95% with inducible MLSB phenotype); mefA, 43% (all with M phenotype); and ermB, 8.5% (45% with inducible MLSB and 45% with constitutive MLSB). Three isolates with constitutive MLSB phenotypes had 23S ribosomal RNA mutations. The 129 erythromycin-resistant isolates belonged to 28 emm types and 44 PFGE patterns, with 51% of the isolates in 4 major PFGE clones each associated with a predominant emm type (emm75, emm58, emm12, and emm114) and resistance genotype (mefA or ermA)). CONCLUSIONS: The population of macrolide-resistant S. pyogenes isolates in the United States is small, but it includes several large clones with potential for expansion.

Otopathogens Detected in Middle Ear Fluid Obtained during Tympanostomy Tube Insertion: Contrasting Purulent and Non-Purulent Effusions.

Otitis media is a prominent disease among children. Previous literature indicates that otitis media is a polymicrobial disease, with Haemophilus influenzae, Streptococcus pneumoniae, Alloiococcus otitidis and Moraxella catarrhalis being the most commonly associated bacterial pathogens. Recent literature suggests that introduction of pneumococcal conjugate vaccines has had an effect on the etiology of otitis media. Using a multiplex PCR procedure, we sought to investigate the presence of the aforementioned bacterial pathogens in middle ear fluid collected from children undergoing routine tympanostomy tube placement at Wake Forest Baptist Medical Center during the period between January 2011 and March 2014. In purulent effusions, one or more bacterial organisms were detected in ~90% of samples. Most often the presence of H. influenzae alone was detected in purulent effusions (32%; 10 of 31). In non-purulent effusions, the most prevalent organism detected was A. otitidis (26%; 63 of 245). Half of the non-purulent effusions had none of these otopathogens detected. In purulent and non-purulent effusions, the overall presence of S. pneumoniae was lower (19%; 6 of 31, and 4%; 9 of 245, respectively) than that of the other pathogens being identified. The ratio of the percentage of each otopathogen identified in purulent vs. non-purulent effusions was >1 for the classic otopathogens but not for A. otitidis.

Srv mediated dispersal of streptococcal biofilms through SpeB is observed in CovRS+ strains.

Group A Streptococcus (GAS) is a human specific pathogen capable of causing both mild infections and severe invasive disease. We and others have shown that GAS is able to form biofilms during infection. That is to say, they form a three-dimensional, surface attached structure consisting of bacteria and a multi-component extracellular matrix. The mechanisms involved in regulation and dispersal of these GAS structures are still unclear. Recently we have reported that in the absence of the transcriptional regulator Srv in the MGAS5005 background, the cysteine protease SpeB is constitutively produced, leading to increased tissue damage and decreased biofilm formation during a subcutaneous infection in a mouse model. This was interesting because MGAS5005 has a naturally occurring mutation that inactivates the sensor kinase domain of the two component regulatory system CovRS. Others have previously shown that strains lacking covS are associated with decreased SpeB production due to CovR repression of speB expression. Thus, our results suggest the inactivation of srv can bypass CovR repression and lead to constitutive SpeB production. We hypothesized that Srv control of SpeB production may be a mechanism to regulate biofilm dispersal and provide a mechanism by which mild infection can transition to severe disease through biofilm dispersal. The question remained however, is this mechanism conserved among GAS strains or restricted to the unique genetic makeup of MGAS5005. Here we show that Srv mediated control of SpeB and biofilm dispersal is conserved in the invasive clinical isolates RGAS053 (serotype M1) and MGAS315 (serotype M3), both of which have covS intact. This work provides additional evidence that Srv regulated control of SpeB may mediate biofilm formation and dispersal in diverse strain backgrounds.