Engineering bacteriocin-mediated resistance against the plant pathogen Pseudomonas syringae.

The plant pathogen, Pseudomonas syringae (Ps), together with related Ps species, infects and attacks a wide range of agronomically important crops, including tomato, kiwifruit, pepper, olive and soybean, causing economic losses. Currently, chemicals and introduced resistance genes are used to protect plants against these pathogens but have limited success and may have adverse environmental impacts. Consequently, there is a pressing need to develop alternative strategies to combat bacterial disease in crops. One such strategy involves using narrow-spectrum protein antibiotics (so-called bacteriocins), which diverse bacteria use to compete against closely related species. Here, we demonstrate that one bacteriocin, putidacin L1 (PL1), can be expressed in an active form at high levels in Arabidopsis and in Nicotiana benthamiana in planta to provide effective resistance against diverse pathovars of Ps. Furthermore, we find that Ps strains that mutate to acquire tolerance to PL1 lose their O-antigen, exhibit reduced motility and still cannot induce disease symptoms in PL1-transgenic Arabidopsis. Our results provide proof-of-principle that the transgene-mediated expression of a bacteriocin in planta can provide effective disease resistance to bacterial pathogens. Thus, the expression of bacteriocins in crops might offer an effective strategy for managing bacterial disease, in the same way that the genetic modification of crops to express insecticidal proteins has proven to be an extremely successful strategy for pest management. Crucially, nearly all genera of bacteria, including many plant pathogenic species, produce bacteriocins, providing an extensive source of these antimicrobial agents.

Selective autophagy limits cauliflower mosaic virus infection by NBR1-mediated targeting of viral capsid protein and particles.

Autophagy plays a paramount role in mammalian antiviral immunity including direct targeting of viruses and their individual components, and many viruses have evolved measures to antagonize or even exploit autophagy mechanisms for the benefit of infection. In plants, however, the functions of autophagy in host immunity and viral pathogenesis are poorly understood. In this study, we have identified both anti- and proviral roles of autophagy in the compatible interaction of cauliflower mosaic virus (CaMV), a double-stranded DNA pararetrovirus, with the model plant Arabidopsis thaliana We show that the autophagy cargo receptor NEIGHBOR OF BRCA1 (NBR1) targets nonassembled and virus particle-forming capsid proteins to mediate their autophagy-dependent degradation, thereby restricting the establishment of CaMV infection. Intriguingly, the CaMV-induced virus factory inclusions seem to protect against autophagic destruction by sequestering capsid proteins and coordinating particle assembly and storage. In addition, we found that virus-triggered autophagy prevents extensive senescence and tissue death of infected plants in a largely NBR1-independent manner. This survival function significantly extends the timespan of virus production, thereby increasing the chances for virus particle acquisition by aphid vectors and CaMV transmission. Together, our results provide evidence for the integration of selective autophagy into plant immunity against viruses and reveal potential viral strategies to evade and adapt autophagic processes for successful pathogenesis.

Child-related attributions of hostile intent and harsh discipline: Moderating effects of anger.

Two studies tested whether attributions of hostile intent (AHI) and anger for ambiguous child behaviors uniquely predict parents' harsh verbal discipline (HVD) and harsh physical discipline (HPD) or whether AHI only predicts harsh discipline when AHI occurs in conjunction with anger, as suggested by the Integrated Cognitive Model of general aggression (Wilkowski & Robinson, 2010, J. Pers., 78, 9-38). Replicating previous studies, the expected positive bivariate relationships between AHI, anger, HVD, and HPD were found in Study 1 (N = 493). However, when AHI, anger, and the AHI by anger interaction were considered simultaneously, patterns of prediction varied by discipline type. For HVD, high levels of AHI, high levels of anger, and the AHI by anger interaction (at high levels of AHI and anger) each were significant predictors. For HPD, low levels of AHI, high levels of anger, and the AHI by anger interaction (at high levels of AHI and anger) were significant predictors, suggesting a possible suppressor effect for AHI. These results were replicated in Study 2 ( N = 503). Thus, there was support for the contention that AHI only predicts harsh discipline when AHI occurs in conjunction with anger with respect to HPD but this was not the case for HVD. Since AHI suppressor effects were observed for HPD in two large sample studies, variables that may account for the AHI suppressor effects and the need for testing more complex interactive predictive models of harsh parenting behaviors are discussed.

Lectin-like bacteriocins from Pseudomonas spp. utilise D-rhamnose containing lipopolysaccharide as a cellular receptor.

Lectin-like bacteriocins consist of tandem monocot mannose-binding domains and display a genus-specific killing activity. Here we show that pyocin L1, a novel member of this family from Pseudomonas aeruginosa, targets susceptible strains of this species through recognition of the common polysaccharide antigen (CPA) of P. aeruginosa lipopolysaccharide that is predominantly a homopolymer of D-rhamnose. Structural and biophysical analyses show that recognition of CPA occurs through the C-terminal carbohydrate-binding domain of pyocin L1 and that this interaction is a prerequisite for bactericidal activity. Further to this, we show that the previously described lectin-like bacteriocin putidacin L1 shows a similar carbohydrate-binding specificity, indicating that oligosaccharides containing D-rhamnose and not D-mannose, as was previously thought, are the physiologically relevant ligands for this group of bacteriocins. The widespread inclusion of d-rhamnose in the lipopolysaccharide of members of the genus Pseudomonas explains the unusual genus-specific activity of the lectin-like bacteriocins.

Stability of small ubiquitin-like modifier (SUMO) proteases OVERLY TOLERANT TO SALT1 and -2 modulates salicylic acid signalling and SUMO1/2 conjugation in Arabidopsis thaliana.

Small ubiquitin-like modifier proteases 1 and 2 (SUMO1/2) have been linked to the regulation of salicylic acid (SA)-mediated defence signalling in Arabidopsis thaliana. In order to define the role of the SUMO proteases OVERLY TOLERANT TO SALT1 and -2 (OTS1/2) in defence and to provide insight into SUMO1/2-mediated regulation of SA signalling, we examined the status of SA-mediated defences in ots1/2 mutants. The ots1 ots2 double mutant displayed enhanced resistance to virulent Pseudomonas syringae and higher levels of SA compared with wild-type (WT) plants. Furthermore, ots1 ots2 mutants exhibited upregulated expression of the SA biosynthesis gene ICS1 in addition to enhanced SA-responsive ICS1 expression beyond that of WT. SA stimulated OTS1/2 degradation and promoted accumulation of SUMO1/2 conjugates. These results indicate that OTS1 and -2 act in a feedback loop in SA signalling and that de novo OTS1/2 synthesis works antagonistically to SA-promoted degradation, adjusting the abundance of OTS1/2 to moderate SA signalling. Accumulation of SUMO1/2 conjugates coincides with SA-promoted OTS degradation and may play a positive role in SA-mediated signalling in addition to its repressive roles reported elsewhere.

Structure of the atypical bacteriocin pectocin M2 implies a novel mechanism of protein uptake.

The colicin-like bacteriocins are potent protein antibiotics that have evolved to efficiently cross the outer membrane of Gram-negative bacteria by parasitizing nutrient uptake systems. We have structurally characterized the colicin M-like bacteriocin, pectocin M2, which is active against strains of Pectobacterium spp. This unusual bacteriocin lacks the intrinsically unstructured translocation domain that usually mediates translocation of these bacteriocins across the outer membrane, containing only a single globular ferredoxin domain connected to its cytotoxic domain by a flexible α-helix, which allows it to adopt two distinct conformations in solution. The ferredoxin domain of pectocin M2 is homologous to plant ferredoxins and allows pectocin M2 to parasitize a system utilized by Pectobacterium to obtain iron during infection of plants. Furthermore, we identify a novel ferredoxin-containing bacteriocin pectocin P, which possesses a cytotoxic domain homologous to lysozyme, illustrating that the ferredoxin domain acts as a generic delivery module for cytotoxic domains in Pectobacterium.

Changes in healthcare use across the transition from civilian to military life.

Patterns of healthcare use in a sample of young adults entering the US Navy (N=1137) were examined in a longitudinal survey study. Baseline data provided information about healthcare use as a civilian, whereas follow-up data were used to examine changes in patterns of use over time following entry into the Military Health System (MHS). Entrance into the MHS was marked by increased use of preventive care. Although few systematic differences were noted with respect to socioeconomic status or race/ethnicity, women consistently used more healthcare than did men, and women's use increased more over time; however, this increase was largely driven by pregnancy during military service. Findings suggest that individuals with access to universal healthcare are likely to increase their overall use of services. However, these effects were quite small in absolute terms, and they were strongest for preventive care rather than more intensive and expensive services. Published 2013. This article is a US Government work and is in the public domain in the USA.

Does parental use of harsh discipline and punishment in response to ambiguous child situations vary by child physical abuse risk?

BACKGROUND/OBJECTIVES: The present study investigated the extent to which fathers and mothers at high-risk and low-risk for child physical abuse (CPA) differed in their use of harsh discipline and punishment when presented ambiguous child situations (situations where differences in positive and negative valences are close to zero). PARTICIPANTS/METHODS: High-risk for CPA parents (N = 74: fathers, n = 41; mothers, n = 33) and low-risk parents (N = 178: fathers, n = 91; mothers, n = 87) were asked to indicate their likelihood of using harsh verbal discipline (HVD), harsh physical discipline (HPD), and punishment (i.e., retributive harm separate from HVD and HPD) after viewing each of eighteen ambiguous child situations. RESULTS: As expected, high-risk, compared to low-risk, for CPA parents were significantly (ps < 0.001) more likely to use HVD (d = 0.546), HPD (d = 0.595), and punishment (d = 0.564). Overall, fathers, relative to mothers, were significantly more likely to use HVD (d = 0.261) and HPD (d = 0.238). For punishment, however, there was no significant parental role (father, mother) difference (d = 0.136). CONCLUSIONS: Since in everyday living situations many child behaviors are ambiguous, the findings that parents, especially high-risk for CPA parents, use harsh discipline and punishment when encountering ambiguous child situations suggest that non-contingent harsh child-related parental behaviors may be frequent in the lives of some children. The findings that parents may use non-contingent discipline and punishment suggest the need to explore whether, when these parenting behaviors occur, they increase the likelihood of negative child outcomes.

The crystal structure of the lipid II-degrading bacteriocin syringacin M suggests unexpected evolutionary relationships between colicin M-like bacteriocins.

Colicin-like bacteriocins show potential as next generation antibiotics with clinical and agricultural applications. Key to these potential applications is their high potency and species specificity that enables a single pathogenic species to be targeted with minimal disturbance of the wider microbial community. Here we present the structure and function of the colicin M-like bacteriocin, syringacin M from Pseudomonas syringae pv. tomato DC3000. Syringacin M kills susceptible cells through a highly specific phosphatase activity that targets lipid II, ultimately inhibiting peptidoglycan synthesis. Comparison of the structures of syringacin M and colicin M reveals that, in addition to the expected similarity between the homologous C-terminal catalytic domains, the receptor binding domains of these proteins, which share no discernible sequence homology, share a striking structural similarity. This indicates that the generation of the novel receptor binding and species specificities of these bacteriocins has been driven by diversifying selection rather than diversifying recombination as suggested previously. Additionally, the structure of syringacin M reveals the presence of an active site calcium ion that is coordinated by a conserved aspartic acid side chain and is essential for catalytic activity. We show that mutation of this residue to alanine inactivates syringacin M and that the metal ion is absent from the structure of the mutant protein. Consistent with the presence of Ca(2+) in the active site, we show that syringacin M activity is supported by Ca(2+), along with Mg(2+) and Mn(2+), and the protein is catalytically inactive in the absence of these ions.

Identification of the domains of cauliflower mosaic virus protein P6 responsible for suppression of RNA silencing and salicylic acid signalling.

Cauliflower mosaic virus (CaMV) encodes a 520 aa polypeptide, P6, which participates in several essential activities in the virus life cycle including suppressing RNA silencing and salicylic acid-responsive defence signalling. We infected Arabidopsis with CaMV mutants containing short in-frame deletions within the P6 ORF. A deletion in the distal end of domain D-I (the N-terminal 112 aa) of P6 did not affect virus replication but compromised symptom development and curtailed the ability to restore GFP fluorescence in a GFP-silenced transgenic Arabidopsis line. A deletion in the minimum transactivator domain was defective in virus replication but retained the capacity to suppress RNA silencing locally. Symptom expression in CaMV-infected plants is apparently linked to the ability to suppress RNA silencing. When transiently co-expressed with tomato bushy stunt virus P19, an elicitor of programmed cell death in Nicotiana tabacum, WT P6 suppressed the hypersensitive response, but three mutants, two with deletions within the distal end of domain D-I and one involving the N-terminal nuclear export signal (NES), were unable to do so. Deleting the N-terminal 20 aa also abolished the suppression of pathogen-associated molecular pattern-dependent PR1a expression following agroinfiltration. However, the two other deletions in domain D-I retained this activity, evidence that the mechanisms underlying these functions are not identical. The D-I domain of P6 when expressed alone failed to suppress either cell death or PR1a expression and is therefore necessary but not sufficient for all three defence suppression activities. Consequently, concerns about the biosafety of genetically modified crops carrying truncated ORFVI sequences appear unfounded.

Does the Fading Affect Bias Vary by Memory Type and a Parent's Risk of Physically Abusing a Child? A Replication and Extension.

Research described in the present article assessed (a) whether a fading affect bias (FAB) occurred in parent memories of a child as well as parent general personal memories and (b) whether either or both of these FAB effects was moderated by a parent's risk of physically abusing a child. A FAB effect, unmoderated by parents' abuse risk status, emerged for parents' general personal memories. In contrast, the FAB was muted when high abuse risk parents remembered child-related events: High abuse risk parents seemed unable to "let go" of negative affect prompted by recall of negative events involving their children. This finding replicates and extends findings reported in prior research. However, this significant moderation effect occurred for only one event memory collection method. This methodological dependence is unusual: the FAB effect is typically impervious to methodological variations. Implications of these results, as well as suggestions for future research, are discussed.

Effect sizes and effect size benchmarks in family violence research.

Scholarly journals increasingly request that authors include effect size (ES) estimates when reporting statistical results. However, there is little guidance on how authors should interpret ESs. Consequently, some authors do not provide ES interpretations, or, when interpretations are provided, they often fail to use appropriate reference groups, using instead the ES benchmarks suggested by Cohen (1988). After discussing the most commonly used ES estimates, we describe the method used by Cohen (1962) to develop ES benchmarks (i.e., small, medium, and large) for use in power analyses and describe the limitations associated with using these benchmarks. Next, we establish general benchmarks for family violence (FV) research. That is, we followed Cohen's approach to establishing his original ES benchmarks using family violence research published in 2021 in Child Abuse & Neglect, which produced a medium ES (d = 0.354) that was smaller than Cohen's recommended medium ES (d = 0.500). Then, we examined the ESs in different subspecialty areas of FV research to provide benchmarks for contextualized FV ESs and to provide information that can be used to conduct power analyses when planning future FV research. Finally, some of the challenges to developing ES benchmarks in any scholarly discipline are discussed. For professionals who are not well informed about ESs, the present review is designed to increase their understanding of ESs and what ES benchmarks tell them (and do not tell them) with respect to understanding the meaningfulness of FV research findings.

Prolonged Infant Crying: Caregiving Quality and Child Physical Abuse Risk.

Research suggests that prolonged infant crying may increase risk for child physical abuse (CPA). However, few studies have examined behavioral responses to infant crying among parents at risk for CPA. The present study sought to fill this gap by using a simulated infant to examine how mothers and fathers with varying degrees of CPA risk respond to prolonged infant crying. Specifically, a sample of 184 general population caregivers (107 mothers and 77 fathers) participated in a task that involved attempting to soothe a simulated infant that cried continuously for 30 min. The simulated infant sessions were video-recorded, and quality of caregiving behavior was coded in 5-min segments. Participants rated their negative affect (feelings of upset, distress, irritability) at the outset of the data collection session, before beginning the simulated infant task, and after the simulated infant task concluded. It was predicted that high-risk caregivers, compared to low-risk caregivers, would display lower quality caregiving, higher levels of negative affect, and that CPA risk group differences would increase over time. Gender differences were explored in relation to the aforementioned hypotheses. Over the course of the 30-min simulated infant task, the quality of caregiving behavior diminished among both high- and low-risk caregivers. As expected, high-risk caregivers, compared to low-risk caregivers, exhibited lower quality caregiving behaviors and were more likely to discontinue the simulated infant task early. In addition, high-risk, compared to low-risk, caregivers reported higher levels of negative affect throughout the data collection session, with the highest level of negative affect reported by high-risk caregivers after completing the simulated infant task. Overall, the quality of caregiving exhibited by men and women did not significantly differ. The present findings highlight the importance of early intervention designed to support caregivers' abilities to respond effectively to prolonged infant crying.

Bacteriocins active against plant pathogenic bacteria.

Gram-negative phytopathogens cause significant losses in a diverse range of economically important crop plants. The effectiveness of traditional countermeasures, such as the breeding and introduction of resistant cultivars, is often limited by the dearth of available sources of genetic resistance. An alternative strategy to reduce loss to specific bacterial phytopathogens is to use narrow-spectrum protein antibiotics such as colicin-like bacteriocins as biocontrol agents. A number of colicin-like bacteriocins active against phytopathogenic bacteria have been described previously as have strategies for their application to biocontrol. In the present paper, we discuss these strategies and our own recent work on the identification and characterization of candidate bacteriocins and how these potent and selective antimicrobial agents can be effectively applied to the control of economically important plant disease.

Do High-Risk Parents Use More Aggression During Competitive Play With Children?

The present study examined the extent to which the aggressive tendencies of parents at risk for perpetrating child physical abuse (CPA) generalize to situations other than discipline-related encounters (e.g., a competitive gaming interaction). Participants included parents who were either low (n = 90) or high (n = 75) risk for CPA. Parents were led to believe that they were playing a game against a child opponent with whom they exchanged sound blasts of varying intensities. Parental sound blast selections served as a proxy for aggressive behavior. Parents were randomly assigned to high or low loss conditions. The fictitious child opponent's responses were pre-programed so that sound blasts received by parents became louder (i.e., more provocative) over time. As predicted, high CPA risk parents exhibited higher levels of aggression than low CPA risk parents. Moreover, high-risk parents reported having higher levels of aggressive motives during the game than low-risk parents; and aggressive motives explained the link between parental CPA risk and parents' aggressive behavior. With one exception, higher levels of child provocation during the game prompted parents to send higher levels of sound blasts. The exception to this pattern occurred among low-risk parents who experienced high rates of loss during the game. Specifically, low-risk parents in the high loss condition continued to send low levels of sound blasts even as their child opponent became more provocative. Aggressive motives (and the ability to change motives as situations change) may help explain differences in aggressive tendencies evinced by parents with varying levels of CPA risk.

Evaluation of a sexual assault education/prevention program for female U.S. Navy personnel.

The U.S. Navy Sexual Assault Intervention Training (SAIT) program for women was evaluated in a randomized clinical trial. The SAIT uses multiple presentation modalities (lecture, slides, discussion, film) to provide information related to sexual assault, including risk factors, consequences, prevention, and relevant military regulations. Female personnel who had completed basic training (N = 550) participated in the SAIT or a Comparison condition, and then completed measures of rape knowledge, empathy for rape victims, and acceptance of rape myths (false beliefs about rape justifying sexual violence). Results showed that the SAIT increased factual knowledge about rape. In addition, the SAIT increased empathy with rape victims in some groups of women. However, the program did not reduce women's rape myth acceptance. Given the enormity of the problem of sexual assault and these promising initial findings, additional research on the efficacy of the SAIT is clearly warranted.

Challenges of using protein antibiotics for pathogen control.

Bacterial phytopathogens represent a significant threat to many economically important crops. Current control measures often inflict harm on the environment and may ultimately impact on human health through the spread of antibiotic resistance. Antimicrobial proteins such as bacteriocins have been suggested as the next generation of disease control agents since they are able to specifically target the pathogen of interest with minimal impact on the wider microbial community and environment. However, substantial gaps in knowledge with regards to the efficacy and application of bacteriocins to combat phytopathogenic bacteria remain. Here we highlight the immediate challenges the community must address to ensure maximum exploitation of antimicrobial proteins in the field. © 2021 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Perceived Executive Functioning in Parents at Risk for Child Physical Abuse.

Research suggests that deficits in executive functioning are associated with negative parenting behaviors. However, limited research has examined the link between executive functioning and risk for child physical abuse (CPA) perpetration. Early studies examining executive functioning in parents at risk for perpetrating CPA relied on performance-based measures, which are designed to occur under carefully controlled conditions and may not capture difficulties experienced under less optimal conditions (e.g., during chaotic caregiving situations). Moreover, prior studies examining executive functioning in parents at risk for perpetrating CPA have relied on small samples comprised of only mothers. To advance our understanding of the linkage between executive functioning and CPA risk, the present study examined perceived deficits in executive functioning in a sample of general population mothers and fathers (N = 98) using a standardized self-report measure of executive functioning, namely, the adult version of the Behavior Rating Inventory of Executive Function (BRIEF-A). Parents were classified as low CPA risk or high CPA risk using the Child Abuse Potential Inventory. Compared with low-risk parents, high-risk parents had higher rates of clinical elevations on several BRIEF-A subscales, namely, Working Memory Problems, Emotional Control Difficulties, and Difficulties Shifting Thoughts, Feelings, and Behaviors. These findings highlight the potential importance of assessing and strengthening executive functioning in interventions designed to reduce risk of parent-to-child aggression.

Evaluation of a sexual assault education/prevention program for male U.S. Navy personnel.

A randomized clinical trial was conducted to evaluate the effectiveness of the Navy Sexual Assault Intervention Training (SAIT) program for men. A four-group Solomon design was used to control for possible pretest sensitization effects. Male Navy personnel (N = 1,505) were assessed for rape knowledge, rape myth acceptance (two scales), and rape empathy after participating in the SAIT program or viewing an educational video about HIV/AIDS (comparison condition). The SAIT program was found to be effective in increasing rape knowledge, reducing rape myth acceptance, and increasing empathy for rape victims. As expected, men who had exhibited previous coercive sexual behavior, compared with those who had not, reported lower levels of knowledge, higher levels of rape myth acceptance, and less rape empathy. However, the SAIT program was generally effective in changing men's knowledge, beliefs, and feelings on the key measures, regardless of participants' histories of coercive sexual behavior.

Substantiation of spouse and child maltreatment reports as a function of referral source and maltreatment type.

Substantiation rates for alleged incidents of spouse (N = 33,787) and child (N = 31,986) maltreatment reported to the U.S. Air Force (AF) Family Advocacy Program between 2000 and 2007 were examined. For spouse maltreatment, physical abuse and multiple forms of maltreatment were most likely to be substantiated and neglect was least likely to be substantiated. For child maltreatment, emotional abuse was most likely to be substantiated and physical abuse was least likely to be substantiated. Substantiation rates were higher for referrals by military professionals than for referrals by civilian professionals or nonprofessionals; considerable variation in substantiation rates within each of these categories also was observed. Overall, AF substantiation rates were higher for spouse than for child maltreatment, and substantiation rates for child maltreatment were higher in the AF than in the general U.S. population. Substantiation rates have declined over time in both the AF and the United States.