Simulation of acoustic reflection and backscatter from arctic sea-ice.

The rapidly warming Arctic ocean demands new ways to monitor and characterize changes in sea-ice distribution, thickness, and mechanical properties. Upward-looking sonars mounted on autonomous underwater vehicles offer possibilities for doing so. Numerical simulations were made of the signal received by an upward-looking sonar under a smooth ice sheet using a wavenumber integration code. Demands on sonar frequency and bandwidth for pulse-echo measurements were analyzed. For typical sea-ice physical properties found in the Arctic ocean, even in highly attenuating sea-ice, there is significant information to be extracted from the received acoustic signal. Discrete resonance frequencies in the signal may be related to leaky Lamb waves, and the frequencies are connected to the ratio of the shear wave speed-to-thickness of the ice sheet. The periodicity of the multiple reflections of a pulse-compressed signal may be related to the ratio of compressional wave speed-to- thickness. Decay rates of both types of signals are indicative of the wave attenuation coefficients. Simulations of the acoustic reflection by rough water-ice interfaces were made. Smaller levels of roughness were found to enhance the acoustic signal, while greater levels of roughness are detrimental to the sea-ice characterization process.

SARS-CoV-2 Testing in the Community: Testing Positive Samples with the TaqMan SARS-CoV-2 Mutation Panel To Find Variants in Real Time.

Genome sequencing is a powerful tool for identifying SARS-CoV-2 variant lineages; however, there can be limitations due to sequence dropout when used to identify specific key mutations. Recently, ThermoFisher Scientific has developed genotyping assays to help bridge the gap between testing capacity and sequencing capability to generate real-time genotyping results based on specific variants. Over a 6-week period during the months of April and May 2021, we set out to assess the ThermoFisher TaqMan mutation panel genotyping assay, initially for three mutations of concern and then for an additional two mutations of concern, against SARS-CoV-2-positive clinical samples and the corresponding COVID-19 Genomics UK Consortium (COG-UK) sequencing data. We demonstrate that genotyping is a powerful in-depth technique for identifying specific mutations, is an excellent complement to genome sequencing, and has real clinical health value potential, allowing laboratories to report and take action on variants of concern much more quickly.

Acoustic propagation in gassy intertidal marine sediments: An experimental study.

The need to predict acoustic propagation through marine sediments that contain gas bubbles has become increasingly important for civil engineering and climate studies. There are relatively few in situ acoustic wave propagation studies of muddy intertidal sediments, in which bubbles of biogenic gas (generally methane, a potent greenhouse gas) are commonly found. We used a single experimental rig to conduct two in situ intertidal acoustical experiments to improve understanding of acoustic remote sensing of gassy sediments, eventually including gas bubble size distributions. In the first experiment, we measured sediment sound speed and attenuation between four aligned hydrophones for a quasi-plane wave propagating along the array. The second experiment involved a focused insonified sediment volume created by two transducers emitting coincident sound beams at different frequencies that generated bubble-mediated acoustic signals at combination frequencies. The results from sediment core analyses, and comparison of in situ acoustic velocity and attenuation values with those of water-saturated sediments, together provide ample evidence for the presence of in situ gas bubbles in the insonified volumes of sediments. These datasets are suitable for linear and non-linear inversion studies that estimate in situ greenhouse gas bubble populations, needed for future acoustical remote sensing applications.

Validation testing to determine the sensitivity of lateral flow testing for asymptomatic SARS-CoV-2 detection in low prevalence settings: Testing frequency and public health messaging is key.

Lateral flow devices (LFDs) are quickly being implemented for use in large-scale population surveillance programs for SARS-CoV-2 infection in the United Kingdom. These programs have been piloted in city-wide screening in the city of Liverpool and are now being rolled out to support care home visits and the return home of University students for the Christmas break. Here, we present data on the performance of LFDs to test almost 8,000 students at the University of Birmingham between December 2 and December 9, 2020. The performance is validated against almost 800 samples using PCR performed in the University Pillar 2 testing lab and theoretically validated on thousands of Pillar 2 PCR testing results performed on low-prevalence care home testing samples. Our data show that LFDs do not detect infections presenting with PCR Ct values over 29 to 30 as determined using the Thermo Fisher TaqPath asssay. This may be of particular importance in detecting individuals that are either at the early, or late stages of infection, and reinforces the need for frequent, recurrent testing.

S-Variant SARS-CoV-2 Lineage B1.1.7 Is Associated With Significantly Higher Viral Load in Samples Tested by TaqPath Polymerase Chain Reaction.

A SARS-CoV-2 variant B1.1.7 containing mutation Δ69/70 has spread rapidly in the United Kingdom and shows an identifiable profile in ThermoFisher TaqPath RT-qPCR, S gene target failure (SGTF). We analyzed recent test data for trends and significance. Linked cycle threshold (Ct) values for respiratory samples showed that a low Ct for ORF1ab and N were clearly associated with SGTF. Significantly more SGTF samples had higher inferred viral loads between 1×107 and 1×108. Our conclusion is that patients whose samples exhibit the SGTF profile are more likely to have high viral loads, which may explain higher infectivity and rapidity of spread.

Geophysical early warning of salt precipitation during geological carbon sequestration.

Sequestration of industrial carbon dioxide (CO2) in deep geological saline aquifers is needed to mitigate global greenhouse gas emissions; monitoring the mechanical integrity of reservoir formations is essential for effective and safe operations. Clogging of fluid transport pathways in rocks from CO2-induced salt precipitation reduces injectivity and potentially compromises the reservoir storage integrity through pore fluid pressure build-up. Here, we show that early warning of salt precipitation can be achieved through geophysical remote sensing. From elastic P- and S-wave velocity and electrical resistivity monitoring during controlled laboratory CO2 injection experiments into brine-saturated quartz-sandstone of high porosity (29%) and permeability (1660 mD), and X-ray CT imaging of pore-scale salt precipitation, we were able to observe, for the first time, how CO2-induced salt precipitation leads to detectable geophysical signatures. We inferred salt-induced rock changes from (i) strain changes, (ii) a permanent ~ 1.5% decrease in wave velocities, linking the geophysical signatures to salt volume fraction through geophysical models, and (iii) increases of porosity (by ~ 6%) and permeability (~ 7%). Despite over 10% salt saturation, no clogging effects were observed, which suggests salt precipitation could extend to large sub-surface regions without loss of CO2 injectivity into high porosity and permeability saline sandstone aquifers.

Identification of a novel prophage regulator in Escherichia coli controlling the expression of type III secretion.

This study has identified horizontally acquired genomic regions of enterohaemorrhagic Escherichia coli O157:H7 that regulate expression of the type III secretion (T3S) system encoded by the locus of enterocyte effacement (LEE). Deletion of O-island 51, a 14.93 kb cryptic prophage (CP-933C), resulted in a reduction in LEE expression and T3S. The deletion also had a reduced capacity to attach to epithelial cells and significantly reduced E. coli O157 excretion levels from sheep. Further characterization of O-island 51 identified a novel positive regulator of the LEE, encoded by ecs1581 in the E. coli O157:H7 strain Sakai genome and present but not annotated in the E. coli strain EDL933 sequence. Functionally important residues of ECs1581 were identified based on phenotypic variants present in sequenced E. coli strains and the regulator was termed RgdR based on a motif demonstrated to be important for stimulation of gene expression. While RgdR activated expression from the LEE1 promoter in the presence or absence of the LEE-encoded regulator (Ler), RgdR stimulation of T3S required ler and Ler autoregulation. RgdR also controlled the expression of other phenotypes, including motility, indicating that this new family of regulators may have a more global role in E. coli gene expression.

Transcriptional regulators of the GAD acid stress island are carried by effector protein-encoding prophages and indirectly control type III secretion in enterohemorrhagic Escherichia coli O157:H7.

Type III secretion (T3S) plays a pivotal role in the colonization of ruminant hosts by Enterohemorrhagic Escherichia coli (EHEC). The T3S system translocates effector proteins into host cells to promote bacterial attachment and persistence. The repertoire and variation in prophage regions underpins differences in the pathogenesis and epidemiology of EHEC strains. In this study, we have used a collection of deletions in cryptic prophages and EHEC O157 O-islands to screen for novel regulators of T3S. Using this approach we have identified a family of homologous AraC-like regulators that indirectly repress T3S. These prophage-encoded secretion regulator genes (psr) are found exclusively on prophages and are associated with effector loci and the T3S activating Pch family of regulators. Transcriptional profiling, mutagenesis and DNA binding studies were used to show that these regulators usurp the conserved GAD acid stress resistance system to regulate T3S by increasing the expression of GadE (YhiE) and YhiF and that this regulation follows attachment to bovine epithelial cells. We further demonstrate that PsrA and effectors encoded within cryptic prophage CP933-N are required for persistence in a ruminant model of colonization.

Gene expression profiles induced by Salmonella infection in resistant and susceptible mice.

Mouse models have been extensively used to investigate the mechanisms of salmonellosis. However, the role of the hosts' local intestinal responses during early stages of infection remain unclear. In this study, transcript array analysis was employed to investigate regulation of gene expression in the murine intestine following oral challenge with Salmonella enterica serovar Enteritidis. Salmonella resistant C3H/HeN mice elicited only weak transcription responses in the ileum even in the presence of bacterial replication and systemic infection. This poor response was surprising given previously published results using in vitro models. Susceptible TLR4-deficient C3H/HeJ mice displayed a stronger response, suggesting a role for TLR4 in dampening the response to Salmonella. Responses of susceptible BALB/c mice were also unremarkable. In contrast, in vitro infection of murine rectal epithelial cells induced a strong transcription response consistent with previous in vitro studies. Although the pattern of genes expressed by the ileal tissue upon in vivo infection were similar in all three mouse lines, the genes up-regulated during in vitro infection were different, indicating that the responses seen in vitro do not mimic those seen in vivo. Taken together these data indicate that in vivo responses to Salmonella, at the level of the intestine, are tightly regulated by the host.

N255 is a key residue for recognition by a monoclonal antibody which protects against Yersinia pestis infection.

Mab7.3 to Yersinia pestis LcrV antigen (LcrV(Ype)) protected J774A.1 macrophages in vitro from killing by a Yersinia pseudotuberculosis strain expressing LcrV(Ype). Of 4 site-directed mutations in the coiled-coil region (148-169) and 7 mutations in the 225-255 sequence of LcrV(Ype), only the mutation of N255 to D255, abrogated the binding of Mab7.3 and reduced its protective capacity against plague. Since the Mab7.3 epitope in LcrV(Ype) (135-275) encompasses a region (136-180) thought to be exposed on the injectisome, we suggest that Mab7.3 protects by binding to LcrV(Ype) and interfering with protein-protein interactions necessary for type three secretion.

Absolute calibration of hydrophones immersed in sandy sediment.

An absolute calibration method has been developed based on the method of three-transducer spherical-wave reciprocity for the calibration of hydrophones when immersed in sandy sediment. The method enables the determination of the magnitude of the free-field voltage receive sensitivity of the hydrophone. Adoption of a co-linear configuration allows the acoustic attenuation within the sediment to be eliminated from the sensitivity calculation. Example calibrations have been performed on two hydrophones inserted into sandy sediment over the frequency range from 10 to 200 kHz. In general, a reduction in sensitivity was observed, with average reductions over the frequency range tested of 3.2 and 3.6 dB with respect to the equivalent water-based calibrations for the two hydrophones tested. Repeated measurements were undertaken to assess the robustness of the method to both the influence of the sediment disturbance associated with the hydrophone insertion and the presence of the central hydrophone. A simple finite element model, developed for one of the hydrophone designs, shows good qualitative agreement with the observed differences from water-based calibrations. The method described in this paper will be of interest to all those undertaking acoustic measurements with hydrophones immersed in sediment where the absolute sensitivity is important.

Escherichia coli O157:H7 colonization in small domestic ruminants.

Enterohaemorrhagic Escherichia coli O157:H7 was first implicated in human disease in the early 1980s, with ruminants cited as the primary reservoirs. Preliminary studies indicated cattle to be the sole source of E. coli O157:H7 outbreaks in humans; however, further epidemiological studies soon demonstrated that E. coli O157:H7 was widespread in other food sources and that a number of transmission routes existed. More recently, small domestic ruminants (sheep and goats) have emerged as important sources of E. coli O157:H7 human infection, particularly with the widespread popularity of petting farms and the increased use of sheep and goat food products, including unpasteurized cheeses. Although the colonization and persistence characteristics of E. coli O157:H7 in the bovine host have been studied intensively, this is not the case for small ruminants. Despite many similarities to the bovine host, the pathobiology of E. coli O157:H7 in small domestic ruminants does appear to differ significantly from that described in cattle. This review aims to critically review the current knowledge regarding colonization and persistence of E. coli O157:H7 in small domestic ruminants, including comparisons with the bovine host where appropriate.

Intermittent Escherichia coli O157:H7 colonisation at the terminal rectum mucosa of conventionally-reared lambs.

In cattle, the lymphoid rich regions of the rectal-anal mucosa at the terminal rectum are the preferred site for Escherichia coli O157:H7 colonisation. All cattle infected by rectal swab administration demonstrate long-term E. coli O157:H7 colonisation, whereas orally challenged cattle do not demonstrate long-term E. coli O157:H7 colonisation in all animals. Oral, but not rectal challenge of sheep with E. coli O157:H7 has been reported, but an exact site for colonisation in sheep is unknown. To determine if E. coli O157:H7 can effectively colonise the ovine terminal rectum, in vitro organ culture (IVOC) was initiated. Albeit sparsely, large, densely packed E. coli O157:H7 micro-colonies were observed on the mucosa of ovine and control bovine terminal rectum explants. After necropsy of orally inoculated lambs, bacterial enumeration of the proximal and distal gastrointestinal tract did suggest a preference for E. coli O157:H7 colonisation at the ovine terminal rectum, albeit for both lymphoid rich and non-lymphoid sites. As reported for cattle, rectal inoculation studies were then conducted to determine if all lambs would demonstrate persistent colonisation at the terminal rectum. After necropsy of E. coli O157:H7 rectally inoculated lambs, most animals were not colonised at gastrointestinal sites proximal to the rectum, however, large densely packed micro-colonies of E. coli O157:H7 were observed on the ovine terminal rectum mucosa. Nevertheless, at the end point of the study (day 14), only one lamb had E. coli O157:H7 micro-colonies associated with the terminal rectum mucosa. A comparison of E. coli O157:H7 shedding yielded a similar pattern of persistence between rectally and orally inoculated lambs. The inability of E. coli O157:H7 to effectively colonise the terminal rectum mucosa of all rectally inoculated sheep in the long term, suggests that E. coli O157:H7 may colonise this site, but less effectively than reported previously for cattle.

A mixture containing galactooligosaccharide, produced by the enzymic activity of Bifidobacterium bifidum, reduces Salmonella enterica serovar Typhimurium infection in mice.

The prebiotic Bimuno is a mixture containing galactooligosaccharide, produced by the galactosyltransferase activity of Bifidobacterium bifidum NCIMB 41171 in the presence of lactose. Previous studies have implicated prebiotics in reducing infections by enteric pathogens, thus it was hypothesized that Bimuno may confer some protection in the murine host from Salmonella enterica serovar Typhimurium (S. Typhimurium) infection. In this study, infection caused by S. Typhimurium SL1344nal(r) in the presence or absence of Bimuno was assessed using tissue culture assays, a murine ligated ileal gut loop model and a murine oral challenge model. In tissue culture adherence and invasion assays with HT-29-16E cells, the presence of approximately 2 mM Bimuno significantly reduced the invasion of S. Typhimurium SL1344nal(r) (P<0.0001). In the murine ligated ileal gut loops, the presence of Bimuno prevented colonization and the associated pathology of S. Typhimurium. In the BALB/c mouse model, the oral delivery of Bimuno prior to challenge with S. Typhimurium resulted in significant reductions in colonization in the five organs sampled, with highly significant reductions being observed in the spleen at 72 and 96 h post-challenge (P=0.0002, <0.0001, respectively). Collectively, the results indicate that Bimuno significantly reduced the colonization and pathology associated with S. Typhimurium infection in a murine model system, possibly by reducing the invasion of the pathogen into host cells.

Role of NleH, a type III secreted effector from attaching and effacing pathogens, in colonization of the bovine, ovine, and murine gut.

The human pathogen enterohemorrhagic Escherichia coli (EHEC) O157:H7 colonizes human and animal gut via formation of attaching and effacing lesions. EHEC strains use a type III secretion system to translocate a battery of effector proteins into the mammalian host cell, which subvert diverse signal transduction pathways implicated in actin dynamics, phagocytosis, and innate immunity. The genomes of sequenced EHEC O157:H7 strains contain two copies of the effector protein gene nleH, which share 49% sequence similarity with the gene for the Shigella effector OspG, recently implicated in inhibition of migration of the transcriptional regulator NF-kappaB to the nucleus. In this study we investigated the role of NleH during EHEC O157:H7 infection of calves and lambs. We found that while EHEC DeltanleH colonized the bovine gut more efficiently than the wild-type strain, in lambs the wild-type strain exhibited a competitive advantage over the mutant during mixed infection. Using the mouse pathogen Citrobacter rodentium, which shares many virulence factors with EHEC O157:H7, including NleH, we observed that the wild-type strain exhibited a competitive advantage over the mutant during mixed infection. We found no measurable differences in T-cell infiltration or hyperplasia in colons of mice inoculated with the wild-type or the nleH mutant strain. Using NF-kappaB reporter mice carrying a transgene containing a luciferase reporter driven by three NF-kappaB response elements, we found that NleH causes an increase in NF-kappaB activity in the colonic mucosa. Consistent with this, we found that the nleH mutant triggered a significantly lower tumor necrosis factor alpha response than the wild-type strain.

Characterization of tccP2 carried by atypical enteropathogenic Escherichia coli.

Atypical enteropathogenic Escherichia coli (EPEC) comprise an important group of paediatric pathogens. Atypical EPEC have reservoirs in farm and domestic animals where they can be either commensal or pathogenic; serogroup O26 is dominant in humans and animals. Central to intestinal colonization by EPEC is the translocation of the type III secretion system effector Tir into enterocytes, which following phosphorylation (Tir-Yp) recruits Nck to activate the N-WASP actin signalling cascade. The authors have recently shown that typical EPEC strains, belonging to the EPEC-2 lineage, carry a tir gene encoding Tir-Yp and can also use the alternative TccP2 actin-signalling cascade. The aim of this study was to determine if tccP2 is found in atypical EPEC isolated from human and farm animals. tccP2 was found at a frequency of 41% in non-O26 EPEC isolates and in 82.3% of the O26 strains. TccP2 of human and animal strains show high level of sequence identity. It is shown that most strains carry a tir gene encoding Tir-Yp. In addition the authors identified two new variants of tir genes in EPEC O104:H12 and NT:H19 strains.

An aflagellate mutant Yersinia enterocolitica biotype 1A strain displays altered invasion of epithelial cells, persistence in macrophages, and cytokine secretion profiles in vitro.

Despite being classically defined as non-pathogenic, there is growing evidence that biotype 1A Yersinia enterocolitica isolates may be aetiological agents of disease in humans. In previous studies, a potential link between motility and the ability of biotype 1A strains to invade cultured epithelial cells was observed. In an attempt to further investigate this finding, a flagella mutant was constructed in a human faecal Y. enterocolitica biotype 1A isolate. The flagella mutation abolished the ability of the strain to invade cultured human epithelial cells, although adherence was not affected. The aflagellate mutant was also attenuated in its ability to survive within cultured macrophages, being cleared after 3 h, whilst the wild-type persisted for 24 h after infection. Examination of cytokine secretion by infected macrophages also suggested that the flagella of biotype 1A strains act as anti-inflammatory agents, decreasing production of tumour necrosis factor (TNF)-alpha whilst increasing secretion of interleukin (IL)-10. Preliminary studies using porcine in vitro organ culture (IVOC) tissue suggested that the flagella mutant was also attenuated in its ability to colonize intestinal tissue.

Environmental regulation and colonization attributes of the long polar fimbriae (LPF) of Escherichia coli O157:H7.

The ability of Escherichia coli O157:H7 to colonize the intestinal epithelia is dependent on the expression of intimin and other adhesins. The chromosome of E. coli O157:H7 carries two loci encoding long polar fimbriae (LPF). These fimbriae mediate adherence to epithelial cells and are associated with colonization of the intestine. In order to increase our knowledge about the conditions controlling their expression and their role in colonization of an animal model, the environmental cues that promote expression of lpf genes and the role of E. coli O157:H7 LPF in intestinal colonization of lambs were investigated. We found that expression of lpf1 was regulated in response to growth phase, osmolarity, and pH; that lpf2 transcription was stimulated during late exponential growth and iron depletion; and that LPF impacts the ability of E. coli O157:H7 to persist in the intestine of infected 6-week-old lambs.

Role of intimin-tir interactions and the tir-cytoskeleton coupling protein in the colonization of calves and lambs by Escherichia coli O157:H7.

Intimin facilitates intestinal colonization by enterohemorrhagic Escherichia coli O157:H7; however, the importance of intimin binding to its translocated receptor (Tir) as opposed to cellular coreceptors is unknown. The intimin-Tir interaction is needed for optimal actin assembly under adherent bacteria in vitro, a process which requires the Tir-cytoskeleton coupling protein (TccP/EspF(U)) in E. coli O157:H7. Here we report that E. coli O157:H7 tir mutants are at least as attenuated as isogenic eae mutants in calves and lambs, implying that the role of intimin in the colonization of reservoir hosts can be explained largely by its binding to Tir. Mutation of tccP uncoupled actin assembly from the intimin-Tir-mediated adherence of E. coli O157:H7 in vitro but did not impair intestinal colonization in calves and lambs, implying that pedestal formation may not be necessary for persistence. However, an E. coli O157:H7 tccP mutant induced typical attaching and effacing lesions in a bovine ligated ileal loop model of infection, suggesting that TccP-independent mechanisms of actin assembly may operate in vivo.

A comparison of Shiga-toxin negative Escherichia coli O157 aflagellate and intimin deficient mutants in porcine in vitro and in vivo models of infection.

Isolation of Shiga-toxin (Stx) positive Escherichia coli O157:H7 from commercially grown pigs has been reported. Furthermore, experimental infection studies have demonstrated that Stx-positive E. coli O157:H7 can persist in 12-week-old experimentally orally inoculated conventional pigs for up to 2 months and that persistence was not dependent upon intimin. We have shown that the flagellum of Stx-negative E. coli O157:H7 does not have a role to play in pathogenesis in ruminant models whereas, in poultry, the flagellum of E. coli O157:H7 was important for long-term persistent infection. The contribution of the flagellum of Stx-negative E. coli O157 in the colonisation of pigs was investigated by adherence assays on a porcine (IPI-21) cell line, porcine in vitro organ culture (IVOC) and experimental oral inoculation of conventional 14-week-old pigs. E. coli O157:H7 NCTC12900nal(r) and isogenic aflagellate and intimin deficient mutants adhered equally well to IPI-21 cells. In porcine IVOC association assays, E. coli O157:H7 NCTC12900nal(r) was associated in significantly higher numbers to tissues from the caecum and the terminal rectum than other sites. The aflagellate and intimin deficient mutants significantly adhered in greater numbers to more IVOC gastrointestinal tissues than the parent. Groups of 14-week-old pigs were dosed orally with 10(10)CFU/10ml of either E. coli O157:H7 NCTC12900nal(r) or isogenic aflagellate and intimin deficient mutants and recovery of each test strain was similar. Histological analysis of pig tissues at post mortem examination revealed that E. coli O157 specifically stained bacteria were associated with the mucosa of the ascending and spiral colon. These data suggest that colonisation and persistence of Stx-negative E. coli O157:H7 in pigs, involves mechanisms that do not require the flagellum or intimin.