Bonds and bytes: The odyssey of structural biology.

Characterizing structural and dynamic properties of proteins and large macromolecular assemblies is crucial to understand the molecular mechanisms underlying biological functions. In the field of structural biology, no single method comprehensively reveals the behavior of biological systems across various spatiotemporal scales. Instead, we have a versatile toolkit of techniques, each contributing a piece to the overall puzzle. Integrative structural biology combines different techniques to create accurate and precise multi-scale models that expand our understanding of complex biological systems. This review outlines recent advancements in computational and experimental methods in structural biology, with special focus on recent Artificial Intelligence techniques, emphasizes integrative approaches that combine different types of data for precise spatiotemporal modeling, and provides an outlook into future directions of this field.

Mammographic features and screening outcome in a randomized controlled trial comparing digital breast tomosynthesis and digital mammography.

PURPOSE: To compare the distribution of mammographic features among women recalled for further assessment after screening with digital breast tomosynthesis (DBT) versus digital mammography (DM), and to assess associations between features and final outcome of the screening, including immunohistochemical subtypes of the tumour. METHODS: This randomized controlled trial was performed in Bergen, Norway, and included 28,749 women, of which 1015 were recalled due to mammographic findings. Mammographic features were classified according to a modified BI-RADS-scale. The distribution were compared using 95 % confidence intervals (CI). RESULTS: Asymmetry was the most common feature of all recalls, 24.3 % (108/444) for DBT and 38.9 % (222/571) for DM. Spiculated mass was most common for breast cancer after screening with DBT (36.8 %, 35/95, 95 %CI: 27.2-47.4) while calcifications (23.0 %, 20/87, 95 %CI: 14.6-33.2) was the most frequent after DM. Among women screened with DBT, 0.13 % (95 %CI: 0.08-0.21) had benign outcome after recall due to indistinct mass while the percentage was 0.28 % (95 %CI: 0.20-0.38) for DM. The distributions were 0.70 % (95 %CI: 0.57-0.85) versus 1.46 % (95 %CI: 1.27-1.67) for asymmetry and 0.24 % (95 %CI: 0.16-0.33) versus 0.54 % (95 %CI: 0.43-0.68) for obscured mass, among women screened with DBT versus DM, respectively. Spiculated mass was the most common feature among women diagnosed with non-luminal A-like cancer after DBT and after DM. CONCLUSIONS: Spiculated mass was the dominant feature for breast cancer among women screened with DBT while calcifications was the most frequent feature for DM. Further studies exploring the clinical relevance of mammographic features visible particularly on DBT are warranted.

Interaction between C-Tb and PPD given concomitantly in a split-body randomised controlled trial.

SETTING: Seven tuberculosis (TB) clinics in South Africa. OBJECTIVE: As both purified protein derivative (PPD) and a Mycobacterium tuberculosis-specific skin test (C-Tb) contain region of difference 1 (RD1) antigens, we conducted a study to evaluate whether there was any interaction between the two during concomitant and separate administration in patients with newly diagnosed culture-positive TB. DESIGN: Adult patients with active TB (n = 456, 20% human immunodeficiency virus infected) were randomised to receive only C-Tb, only PPD, or concomitant injection of both C-Tb and PPD using the Mantoux technique. Indurations were read after 48-72 h. QuantiFERON®-TB Gold In-Tube (QFT) was performed in tandem. RESULTS: Of the 456 study participants, 154 simultaneously received both C-Tb and PPD, 153 only C-Tb and 149 only PPD. There was no effect of concomitant injection of PPD on the mean C-Tb induration (19 mm, 95%CI 17-22 vs. 18 mm, 95%CI 16-21; P = 0.91). In patients with active TB, C-Tb sensitivity (78%) was similar to PPD (81%) and QFT (84%; excluding 82/429 [19%] indeterminate results). All tests showed reduced sensitivity in participants with CD4 <100 cells/µl. CONCLUSION: In patients with active TB, there was no interaction between C-Tb and PPD during concomitant injection of both agents. Sensitivities were similar for PPD and C-Tb.

The Efficiency of Biofilters at Mitigating Airborne MRSA from a Swine Nursery.

Our prior studies have been in agreement with other researchers in detecting airborne methicillin-resistant Staphylococcus aureus (MRSA) inside and downwind of a swine housing facility. MRSA emitted in the exhaust air of swine facilities creates a potential risk of transmission of these organisms to people in the general area of these facilities as well as to other animals. This study investigated a possible means of reducing those risks. We investigated the efficiency of biofilters to remove MRSA from the exhaust air of a swine building. Two types of biofilter media (hardwood chips and western red cedar shredded bark) were evaluated. Efficiency was measured by assessing both viable MRSA (viable cascade impactor) and dust particles (optical particle courter) in the pre-filtered and post-filtered air of a functioning swine production facility. Our study revealed that hardwood chips were respectively 92% and 88% efficient in removing viable MRSA and total dust particles. Western red cedar was 95% efficient in removing viable MRSA and 86% efficient in removing dust particles. Our findings suggest that biofilters can be used as effective engineering controls to mitigate the transmission of aerosolized MRSA in the exhaust air of enclosed swine housing facilities.

The effectiveness of BCG vaccination in preventing Mycobacterium tuberculosis infection and disease in Greenland.

BACKGROUND: The BCG vaccine's ability to prevent Mycobacterium tuberculosis infection (MTI) remains highly debated. In Greenland, BCG vaccination was introduced in 1955, but was temporarily discontinued (1991-1996) due to nationwide policy changes. The study aimed to use the transient stop in BCG vaccination to evaluate the effect of vaccination on MTI prevalence and TB incidence. METHODS: MTI study: A cross-sectional study (2012), comprising East Greenlanders born during 1982-2006, evaluated the effect of BCG vaccination on MTI prevalence; a positive interferon γ release assay defined an MTI case. Associations were estimated using logistic regression. TB study: a cohort study covering the same birth cohorts with follow-up until 2012 evaluated the vaccine's effect on TB incidence. A personal identifier allowed for follow-up in the TB notification system. Associations were estimated using Cox regression. RESULTS: MTI study: Included 953 participants; 81% were BCG-vaccinated; 29% had MTI, 23% among vaccinated and 57% among non-vaccinated. BCG vaccination reduced the odds of MTI, OR 0.52 (95% CI 0.32 to 0.85), p=0.01. Vaccine effectiveness against MTI was 20%. TB study: Included 1697 participants followed for 21,148 person-years. 6% were notified with TB, 4% among vaccinated and 11% among non-vaccinated. BCG vaccination reduced the risk of TB, HR 0.50 (95% CI 0.26 to 0.95), p=0.03, yielding a vaccine effectiveness of 50%. CONCLUSIONS: BCG vaccination was effective in reducing both MTI and TB disease among children and young adults in a TB high-endemic setting in Greenland.

Using classification trees to detect induced sow lameness with a transient model.

Feet and legs issues are some of the main causes for sow removal in the US swine industry. More timely lameness detection among breeding herd females will allow better treatment decisions and outcomes. Producers will be able to treat lame females before the problem becomes too severe and cull females while they still have salvage value. The objective of this study was to compare the predictive abilities and accuracies of weight distribution and gait measures relative to each other and to a visual lameness detection method when detecting induced lameness among multiparous sows. Developing an objective lameness diagnosis algorithm will benefit animals, producers and scientists in timely and effective identification of lame individuals as well as aid producers in their efforts to decrease herd lameness by selecting animals that are less prone to become lame. In the early stages of lameness, weight distribution and gait are impacted. Lameness was chemically induced for a short time period in 24 multiparous sows and their weight distribution and walking gait were measured in the days following lameness induction. A linear mixed model was used to determine differences between measurements collected from day to day. Using a classification tree analysis, it was determined that the mean weight being placed on each leg was the most predictive measurement when determining whether the leg was sound or lame. The classification tree's predictive ability decreased as the number of days post-lameness induction increased. The weight distribution measurements had a greater predictive ability compared with the gait measurements. The error rates associated with the weight distribution trees were 29.2% and 31.3% at 6 days post-lameness induction for front and rear injected feet, respectively. For the gait classification trees, the error rates were 60.9% and 29.8% at 6 days post-lameness induction for front and rear injected feet, respectively. More timely lameness detection can improve sow lifetime productivity as well as animal welfare.

Validation of a lameness model in sows using physiological and mechanical measurements.

The objective of this study was to develop a validated, transient, chemically induced lameness model in sows using subjective and objective lameness detection tools. Experiment 1 determined an effective joint injection technique based on volume and placement of dye using feet collected from 9 finisher pigs and 10 multiparity cull sow carcasses. Experiment 2 confirmed the injection technique in live animals and produced a transient clinical lameness in 4 anesthetized sows injected with amphotericin B (15 mg/mL) in the distal interphalangeal joints of the claw. Clinical lameness was assessed by a categorical lameness scoring system, and a postmortem visual confirmation of joint injection technique was obtained. In Exp. 3, 6 sows were injected with 0, 10, or 15 mg/mL amphotericin B in either the left or right hind foot and were monitored until clinical resolution. Treated sows demonstrated elevated clinical lameness scores. These changes resolved by 7 d after lameness induction. Control sows injected with sterile saline developed a clinical lameness score of 0.5, which resolved 72 h post injection. In Exp. 4, 36 sows were injected with 10 mg/mL amphotericin B in 1 of 4 injection sites (left front claws, right front claws, left rear claws, and right rear claws). All injected sows exhibited a decrease in maximum pressure, stance time, and number of sensors activated on the GaitFour (P < 0.05) sensor system. A static force plate also demonstrated a decrease in weight (kg) being placed on the injected foot when all feet were injected (P ≤ 0.05). Injection of amphotericin B induced a predictable acute lameness that resolved spontaneously and is an effective method to model lameness in sows.

Kinetics of UV(254) inactivation of selected viral pathogens in a static system.

AIMS: The objective of this study was to estimate UV(254) inactivation constants for four viral pathogens: influenza virus type A, porcine respiratory and reproductive syndrome virus (PRRSV), bovine viral diarrhoea virus (BVDV) and reovirus. METHODS AND RESULTS: Viruses in culture medium were exposed to one of nine doses of UV(254) and then titrated for infectious virus. Analysis showed that viral inactivation by UV(254) was more accurately described by a two-stage inactivation model vs a standard one-stage inactivation model. CONCLUSIONS: The results provided evidence for the existence of two heterogeneous viral subpopulations among the viruses tested, one highly susceptible to UV(254) inactivation and the other more resistant. Importantly, inactivation constants based on the one-stage inactivation model would have underestimated the UV(254) dose required for the inactivation of these viruses under the conditions of the experiment. SIGNIFICANCE AND IMPACT OF THE STUDY: To improve the accuracy of estimates, it is recommended that research involving the inactivation of micro-organisms evaluates inactivation kinetics using both one-stage and two-stage models. These results will be of interest to persons responsible for microbial agents under laboratory or field conditions.

Using the systematic review methodology to evaluate factors that influence the persistence of influenza virus in environmental matrices.

Understanding factors that influence persistence of influenza virus in an environment without host animals is critical to appropriate decision-making for issues such as quarantine downtimes, setback distances, and eradication programs in livestock production systems. This systematic review identifies literature describing persistence of influenza virus in environmental samples, i.e., air, water, soil, feces, and fomites. An electronic search of PubMed, CAB, AGRICOLA, Biosis, and Compendex was performed, and citation relevance was determined according to the aim of the review. Quality assessment of relevant studies was performed using criteria from experts in virology, disease ecology, and environmental science. A total of 9,760 abstracts were evaluated, and 40 appeared to report the persistence of influenza virus in environmental samples. Evaluation of full texts revealed that 19 of the 40 studies were suitable for review, as they described virus concentration measured at multiple sampling times, with viruses detectable at least twice. Seven studies reported persistence in air (six published before 1970), seven in water (five published after 1990), two in feces, and three on surfaces. All three fomite and five air studies addressed human influenza virus, and all water and feces studies pertained to avian influenza virus. Outcome measurements were transformed to half-lives, and resultant multivariate mixed linear regression models identified influenza virus surviving longer in water than in air. Temperature was a significant predictor of persistence over all matrices. Salinity and pH were significant predictors of persistence in water conditions. An assessment of the methodological quality review of the included studies revealed significant gaps in reporting critical aspects of study design.

Rapid aerosol transmission of Salmonella among turkeys in a simulated holding-shed environment.

Once turkeys arrive at Midwest processing plants, they are usually held in large open-sided sheds for 1 to 4 h, waiting to be unloaded. In hot, humid weather, large fans are used to cool the birds. The resultant air currents distribute a significant amount of dust to the turkeys. The dust created in this environment could be a factor in the number of Salmonella-contaminated turkeys entering slaughter plants. The objective of this study was to determine if rapid transmission of Salmonella in turkeys could occur from exposure to Salmonella-contaminated dust similar to what may be experienced in holding sheds or in other high-dust environments prior to slaughter. In the first experiment, trials of 3 different concentrations of Salmonella (1.2 x 10(9), 2.6 x 10(7), and 2.6 x 10(5) cfu/g) were conducted to determine if transmission of Salmonella enterica serovar Typhimurium var. typhimurium(chi)4232 to turkeys 2 to 4 h after aerosol exposure to contaminated feces is possible. Results showed that turkeys became infected after 2 h of exposure to airborne-contaminated feces with a concentration level of 2.6 x 10(5) cfu of Salmonella Typhimurium/g. In the second experiment, consisting of 3 trials, 1 bank (5 cages wide and 3 cages high) of turkeys (n = 15 birds per trial) was exposed to another bank of cages of S. Typhimurium-inoculated (n = 15) birds for 2 to 4 h using a fan similar to the type in processing-plant cooling sheds. Results from this experiment demonstrated that birds could be contaminated with S. Typhimurium after 2 h of exposure. Results of both studies implicate contaminated dust as a route of rapid airborne transmission of Salmonella in turkeys. Processes that generate significant dust prior to slaughter should be regarded as critical control points for Salmonella.

Optimization of a sampling system for recovery and detection of airborne porcine reproductive and respiratory syndrome virus and swine influenza virus.

The objective of this research was to optimize sampling parameters for increased recovery and detection of airborne porcine reproductive and respiratory syndrome virus (PRRSV) and swine influenza virus (SIV). Collection media containing antifoams, activated carbons, protectants, and ethylene glycol were evaluated for direct effects on factors impacting the detection of PRRSV and SIV, including virus infectivity, viability of continuous cell lines used for the isolation of these viruses, and performance of reverse transcriptase PCR assays. The results showed that specific compounds influenced the likelihood of detecting PRRSV and SIV in collection medium. A subsequent study evaluated the effects of collection medium, impinger model, and sampling time on the recovery of aerosolized PRRSV using a method for making direct comparisons of up to six treatments simultaneously. The results demonstrated that various components in air-sampling systems, including collection medium, impinger model, and sampling time, independently influenced the recovery and detection of PRRSV and/or SIV. Interestingly, it was demonstrated that a 20% solution of ethylene glycol collected the greatest quantity of aerosolized PRRSV, which suggests the possibility of sampling at temperatures below freezing. Based on the results of these experiments, it is recommended that air-sampling systems be optimized for the target pathogen(s) and that recovery/detection results should be interpreted in the context of the actual performance of the system.

The occurrence of asymptomatic and symptomatic simple hepatic cysts. A prospective, hospital-based study.

AIM: To examine the prevalence of asymptomatic and symptomatic liver cysts in a university hospital patient population using modern US equipment. METHODS: Abdominal US scans of 1541 cases referred during the period 21 January to 11 November 2000 were examined for hepatic cysts. RESULTS: Of 1541 cases, 174 (11.3%) were found to have hepatic cysts, i.e. 109 female (12.5%) and 65 (9.7%) male patients (9.7%). In 413 individuals younger than 40 years, no cysts were found. Above the age of 40 years, prevalence increased with age. CONCLUSION: By using modern US equipment, we found a higher prevalence of hepatic cysts than that reported in previous studies. Patient selection and the prevalence of liver cysts in the population from which the patients were referred may have influenced our results.

Allergic sensitisation and allergic rhinitis are associated with n-3 polyunsaturated fatty acids in the diet and in red blood cell membranes.

OBJECTIVE: Due to inconsistent results based on dietary intake data, unsaturated fatty acids in red blood cell (RBC) membranes and diet were used to investigate their association with allergic sensitisation and allergic rhinitis. DESIGN: Cross-sectional, population-based study. SETTING: Bavarian Nutrition Survey II (2002-03), Germany. SUBJECTS: A total of 568 adult participants, 325 women and 243 men. METHODS: By means of logistic regression models, the relation of fatty acids to (i) allergic sensitisation as defined by means of specific serum immunoglobulin E analysis (CAPSX1 class > or = 2), and (ii) self-reported allergic rhinitis was examined. RESULTS: A high cell membrane level of eicosapentaenoic acid (EPA, 20:5 n-3) was inversely associated with allergic sensitisation, the adjusted odds ratio (OR) and 95% confidence interval (95% CI) were 0.52 (0.30-0.90) for the highest (vs lowest) quartile. A similar effect was observed for allergic rhinitis with an OR (95% CI) of 0.50 (0.24-1.03; P = 0.027 for trend). A higher dietary intake of alpha-linolenic acid (ALA, 18:3 n-3) was associated with a decreased risk of allergic sensitisation and allergic rhinitis with ORs (95% CIs) of 0.51 (0.28-0.93) and 0.43 (0.20-0.93), respectively, in the highest quartiles. No other dietary or cell membrane unsaturated fatty acid was significantly associated with the outcome variables, nor was the n-6/n-3 ratio. The strongest effects were observed among subjects under the age of 40 y. CONCLUSIONS: In this cross-sectional study among adults, a high content of n-3 fatty acids in RBC membranes (EPA) or in the diet (ALA) is associated with a decreased risk of allergic sensitisation and allergic rhinitis.

Cytokine and IL-12 receptor mRNA discriminate between different clinical subtypes in multiple sclerosis.

Little is known about the involvement of cytokines in the pathogenesis of primary progressive (PP) multiple sclerosis (MS). We evaluated in this cross-sectional study whether IL-18, IL-12p35, IL-12p40, TNF-alpha, IFN-gamma, IL-10, IL-4, TGF-beta, IL-12Rbeta1, and IL-12Rbeta2 mRNA expression in unstimulated white blood cells showed significant differences between relapsing-remitting (RR), secondary progressive (SP) and PP MS patients, and healthy controls. All clinical subtypes showed unique mRNA expression patterns as compared to the controls. Both RR and SP patients displayed increased levels of IL-12p40, IL-18, and TGF-beta mRNA compared to controls, whereas PP patients showed only increased IL-18 mRNA levels. Both in PP and SP patients, IFN-gamma and IL-10 mRNA were decreased compared to RR patients and controls. PP patients were unique in that they showed decreased IL-12Rbeta1 mRNA. In conclusion, our data show that the assessment of cytokine (receptor) mRNA profiles is useful to discriminate between the different clinical subtypes and suggest that different cytokines are involved in the pathogenesis of PP MS as compared to RR and SP MS.

Contrasting responses to interferon beta-1b treatment in relapsing-remitting multiple sclerosis: does baseline interleukin-12p35 messenger RNA predict the efficacy of treatment?

Interferon (IFN)-beta treatment is effective in relapsing-remitting multiple sclerosis (RR-MS) via an as yet unidentified mechanism. In the present study, we investigated whether the expression of messenger RNA (mRNA) encoding the interleukin (IL)-12 subunits p40 and p35, IL-12 receptor chains, IL-18, tumor necrosis factor-alpha (TNFalpha), IFNgamma, IL-10, IL-4, or transforming growth factor-beta in unstimulated whole blood of 26 RR-MS patients changed during 6 months of IFNbeta-1b treatment. In these patients, a significant change was found in TNFalpha mRNA, whereas changes in IL-12 receptor-beta2 and IL-10 mRNA showed a trend. IFNbeta-1b-related changes in cytokine mRNA expression were next evaluated in clinical subgroups of RR-MS patients classified as either clinical responders or nonresponders on the basis of Expanded Disability Status Scale progression and the number of relapses and steroid interventions needed in the 2 years before initiation of treatment compared with the 2 years after initiation of treatment. These subgroups showed different response patterns to IFNbeta-1b treatment with respect to IL-10, TNFalpha, and IL-18 only. Surprisingly, clinical responders displayed no change in these cytokines, whereas nonresponders showed a decrease in TNFalpha and IL-18 mRNA as well as a transient increase in IL-10 mRNA. Baseline levels of IL-12p35 mRNA were lower in the responders compared with the nonresponders: this marker correctly predicted the clinical outcome in 81% of the 26 patients under investigation.

Using olfactometry to measure intensity and threshold dilution ratio for evaluating swine odor.

Intensity and threshold dilution ratio are two important indices for odor control of swine buildings. Although odor threshold dilution ratio is a widely used index to describe an odor, it should be related to intensity to be more useful. A method was proposed to measure both indices simultaneously by using a dynamic forced-choice olfactometer. Four air samples were taken from each of four swine rooms including farrowing, finisher, gestation, and nursery. A panel of eight people was used to evaluate odor intensity. Odor threshold dilution ratios were calculated according to the American Society for Testing and Materials (ASTM) Standard Practice E679-91 to be 333, 424, 25, and 221 for samples collected from farrowing, finisher, gestation, and nursery rooms, respectively. After the samples were diluted 14.7 times, the odor intensities were evaluated to be 3.79, 3.46, 0.48, and 4.0 for the above-mentioned rooms, respectively. The data collected were used to develop a mathematical model.

Decreased interleukin-10 and increased interleukin-12p40 mRNA are associated with disease activity and characterize different disease stages in multiple sclerosis.

It has been shown that proinflammatory and antiinflammatory cytokines correlate with disease activity in multiple sclerosis (MS). To establish whether such correlations depend on the disease stage, we assessed in a longitudinal fashion the expression of interleukin (IL)-12 (p40 and p35), tumor necrosis factor-alpha, interferon-gamma, and IL-10 mRNA by competitive polymerase chain reaction in unstimulated peripheral blood mononuclear cells of relapsing-remitting (RR) and secondary progressive (SP) MS patients, in relation to monthly clinical and magnetic resonance imaging monitoring. MS patients had increased levels of IL-12p40 and decreased levels of IL-10 mRNA compared with controls; this difference was most pronounced in SP patients. Both RR and SP patients had increased levels of IL-12p40 mRNA compared with controls during the development of active lesions. Moreover, in RR MS an increase was found before relapse. IL-12p35 mRNA was decreased in both groups, and in relation to disease activity it showed a pattern different from IL-12p40 mRNA. In RR MS, IL-10 mRNA was low 4 weeks before magnetic resonance imaging activity and 6 weeks before relapse; a significant increase to normal levels was noted when active lesions became apparent. In contrast, SP patients showed low IL-10 mRNA levels constitutively, suggesting that IL-10 plays an important role in the control of disease progression.

Breakage and deformation characteristics of hypodermic devices under static and dynamic loading.

OBJECTIVE: To characterize the strength and limitations of hypodermic needles to reduce the risk of leaving broken needles in the flesh of animals. SAMPLE POPULATION: Skin of porcine cadavers. Procedure-Stainless steel needles of various gauges combined with aluminum and plastic hubs were subjected to standard test-stand conditions to compare strength under various loading regimens. A device that simulated animal motion was constructed to test breakage characteristics during animal movement. RESULTS: Needles and needle/hub assemblies were resilient to needle breakage, except when bent needles were straightened and a load reapplied. Needle gauge and length drastically affected strength. For 16-and 20-gauge needles, a 1.0-in needle was 1.6 times stronger than a 1.5-in needle. Adding animal movement for 20-gauge, 1.5-in needles resulted in a 40% increase in hub failures for plastic, compared to aluminum hub needles. CONCLUSIONS AND CLINICAL RELEVANCE: Findings of this study are important considerations for meat packers to address in their Hazard Analysis and Critical Control Points plans.