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Ersen B Colkesen1,2, Bart S Ferket2,3, Jan GP Tijssen1, Roderik A Kraaijenhagen2, Coenraad K van Kalken2, Ron JG Peters11Department of Cardiology, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands; 2NDDO Institute for Prevention and Early Diagnostics (NIPED), Amsterdam, the Netherlands; 3Department of Epidemiology, Erasmus Medical Center, Rotterdam, the NetherlandsIntroduction: A large proportion of the cardiovascular disease (CVD) burden can potentially be prevented by primary prevention programs addressing major causal risk factors. A Web-based health risk assessment (HRA) with tailored feedback for individual health promotion is a promising strategy. We evaluated the effect on CVD risk of such a program among employees of a Dutch worksite.Methods: We conducted a prospective follow-up study among 368 employees who voluntarily participated in a Web-based HRA program at a single Dutch worksite in 2008. The program included a multicomponent HRA through a Web-based electronic questionnaire, biometrics, and laboratory evaluation. The results were combined with health behavior change theory to generate tailored motivational and educational health advice. On request, a health counseling session with the program physician was available. Follow-up data on CVD risk were collected 1 year after initial participation. The primary outcome was a change in Framingham CVD risk at 6 months relative to baseline. We checked for a possible background effect of an increased health consciousness as a consequence of program introduction at the worksite by comparing baseline measurements of early program participants with baseline measurements of participants who completed the program 6 months later.Results: A total of 176 employees completed follow-up measurements after a mean of 7 months. There was a graded relation between CVD risk changes and baseline risk, with a relative reduction of 17.9% (P = 0.001) in the high-risk category (baseline CVD risk ≥20%). Changes were not explained by additional health counseling, medication, or an increase in health consciousness within the company.Conclusions: Voluntary participation in a Web-based HRA with tailored feedback at the worksite reduced CVD risk by nearly 18% among participants at high CVD risk and by nearly 5% among all participants. Web-based HRA could improve CVD risk in similar populations. Future research should focus on the persistence of the effects underlying the CVD risk reduction.Keywords: primary prevention, health risk assessment, cardiovascular disease risk

It is not known how membrane fusion proteins that function at neutral pH, for example the human immunodeficiency virus envelope (Env) glycoprotein and intracellular fusion machines, are activated for target bilayer binding. We have addressed this question using a soluble oligomeric form of an avian retroviral Env glycoprotein (API) and soluble forms of its receptor. Binding of soluble receptor to API induces API to bind to liposomes composed of phosphatidylcholine and cholesterol at neutral pH. Liposome binding only occurs at fusion permissive temperatures (T > 20 degrees C), is complete between 2 to 5 min at 37 degrees C, and is stable to high salt, carbonate, and urea. Liposome binding is mediated by the ectodomain of the transmembrane subunit of API, and a mutant with a Val to Glu substitution in the Env fusion peptide (located in the ectodomain of the transmembrane subunit) shows significantly reduced liposome binding. Moreover, under conditions of equivalent binding to API, a mutant receptor that does not support infection (Zingler, K., and J.A.T. Young. 1996. J. Virol. 70:7510-7516) does not induce significant liposome binding. Our results indicate that a highly specific interaction between an avian retroviral Env and its receptor activates the retroviral glycoprotein for target bilayer binding at neutral pH in much the same way as low pH activates the influenza hemagglutinin. Our findings are discussed in terms of the mechanisms of viral and cellular fusion proteins that function at neutral pH.

OBJECTIVE: To describe patients with a distinct electrocardiogram (ECG) pattern without ST-segment elevation in the presence of an acute occlusion of the proximal left anterior descending (LAD) artery. DESIGN: Single-centre observational study. PATIENTS: Patients with acute anterior wall myocardial infarction who were referred for primary percutaneous coronary intervention (PCI) between 1998 and 2008. RESULTS: We identified patients with a static, distinct ECG pattern without ST-segment elevation and an occlusion of the proximal LAD artery during urgent coronary angiography before PCI. Of 1890 patients who underwent primary PCI of the LAD artery, we could identify 35 patients (2%) with this distinct ECG pattern. The ECG showed ST-segment depression at the J-point of at least 1 mm in precordial leads with upsloping ST-segments continuing into tall, symmetrical T-waves. Patients with this distinct ECG pattern were younger, more often male and more often had hypercholesterolaemia compared to patients with anterior myocardial infarction and ST-segment elevation. CONCLUSIONS: In patients presenting with chest pain, ST-segment depression at the J-point with upsloping ST-segments and tall, symmetrical T-waves in the precordial leads of the 12-lead ECG signifies proximal LAD artery occlusion. It is important for cardiologists and emergency care physicians to recognise this distinct ECG pattern, so they can triage such patients for immediate reperfusion therapy. [ABSTRACT FROM AUTHOR]

Statistics show that the prevalence of crack cocaine use and embalming fluid and phencyclidine (PCP)-laced cigarettes or marijuana sticks, commonly referred to on the street as 'fry' or 'wet' is a problem; however, the relationship between these substances of abuse and concurrent polydrug use is unknown. In the present study, a cross-sectional survey was conducted among 426 African-American crack users in Houston, Texas, to investigate the difference between those who concurrently reported lifetime (defined as at least one usage of fry in life) fry use and those who stated they never used fry. The data were analyzed using chi-square and logistic regression analyses. Fry users were significantly more likely than non-users to not have a casual sex partner (92% users vs. 84% non-users, p <= 0.05) and were more likely to have been diagnosed with gonorrhea in the past 12 months (9% users vs. 2% non-users, p <= 0.05). In addition fry users had significantly higher odds of currently trading sex for drugs (OR = 2.30, p <= 0.05), marijuana use (OR = 12.11, p <= 0.05), and codeine (syrup) use (OR = 8.10, p <= 0.05). These findings are important in determining the 'cultural novelties' relative to crack and fry use among younger African Americans. [ABSTRACT FROM AUTHOR]

Despite the availability of the female condoms and theoretically based interventions to promote its use, studies have indicated a low level of acceptability of their use among women in most populations. We aimed to determine female condom use prevalence and the potential markers among African-American women. In an intervention trial to test the efficacy of the Information-Motivation-Behavioral Skills model in increasing condom use, we utilized the baseline data of 280 subjects and examined the potential predictors of female condom use. Chi square statistic and unconditional logistic regression were used to test for group independence among users and non-users of the female condom and to assess the potential markers of female condom use respectively. After adjustment for relevant covariates associated independently with female condom use, the significant potential markers for female condom use were age, multiple sexual relationships, knowledge of female condom, and educational status. Women having multiple sexual relationships compared with a monogamous relationship were five times more likely to use the female condom, while women with high school education were three times more likely to use the female condom; prevalence odds ratio, POR=5.32, 95% CI=1.79-15.83 and POR=3.01, 95% CI=1.01-8.93. Women who were not knowledgeable of the female condom, compared to those who were, were 81% less likely to use the female condom, POR=0.19, 95% CI=0.08-0.45. Among African-American women in this sample, knowledge of female condom use, age, educational status, and multiple sexual relationships were significant markers of female condom use. This study is therefore suggestive of the need to educate African-American women on female condom use, given the obstacles in male condom negotiation, especially among the socio-economically challenged. [ABSTRACT FROM AUTHOR]

While the epidemiologic trends concerning alprazolam (Xanax(R)) are unknown, the use of benzodiazepines, in general, has increased in popularity among youth within recent years. To shed light on the drug problem, the current pilot study used a qualitative approach to investigate relevant beliefs, norms, and perceived addiction associated with alprazolam initiation among 46 youth who were attending an inpatient drug treatment program during the spring of 2004. Overwhelmingly, most participants stated that addiction to alprazolam occurs as early as initial consumption. Most youth in the study stated that their friends felt it was normal to use alprazolam. In addition, their control beliefs revealed that if someone wanted to stop it would be difficult because of the widespread use in their communities and family social reinforcement involved with its use. In this study, a majority of students stated that medical professionals such as doctors and pharmacists were the greatest facilitator of alprazolam acquisition. Implications for these results are discussed. [ABSTRACT FROM AUTHOR]

Raves are characterized by large numbers of youth dancing for long periods of time and by the use of 'club drugs,' such as 3, 4-methylenedioxymethamphetamine (MDMA or 'ecstasy'). While a small body of research has explored the use of ecstasy and other club drugs (EOCD) among club rave attendees in the United States, we are aware of no studies that have investigated the relationship between EOCD use and high-risk sexual behaviors among members of this population. We explored the association between EOCD use and high-risk sexual behaviors among a sample of 283 club rave attendees interviewed during the spring and summer of 2005. Data were collected at 13 rave events in two different clubs along the Baltimore- Washington corridor between March 17 and September 3, 2005. Potential respondents were conveniently sampled and approached between 12 a.m. and 5 a.m. as they exited the clubs. Only 12% reported using ecstasy within the two days preceding the interview, findings considerably lower than our earlier studies of club rave attendees. Moreover, no significant relationship was discerned between recent MDMA use and high-risk sexual behavior. These findings suggest that the use of EOCD among rave attendees has diminished and that the allocation of significant resources to combat their proliferation, among this and other populations at risk for EOCD use, may be premature. [ABSTRACT FROM AUTHOR]

Data on self-reported perceived exposure to anti-smoking messages were collected from 1,608 high school students surveyed through the ASPIRE (A Smoking Prevention Interactive Experience) Program in Houston, Texas. Data collection took place between October 2002 and March 2003. Logistic regression identified that African Americans perceived significantly less exposure to anti-smoking advertisements via television (OR = .50, p 0.05) and posters (OR = .61, p 0.05) than whites. However, they had nearly twice as much perceived exposure to anti-smoking advertisements at movies (OR = 1.79, p 0.05) and sporting events (OR = 2.1, p 0.05) than their white counterparts. Hispanic youth perceived significantly less exposure to anti-smoking posters (OR = .51, p 0.05) and significantly higher exposure to anti-smoking messages at sporting events (OR = 1.92, p 0.05) and school programs (OR = 3.44, p 0.05) compared to white youth. While the relationships tested in this study are exploratory, they provide initial evidence for an important nexus between race and communication channels that may impact perceived exposure to tobacco advertising. [ABSTRACT FROM AUTHOR]

Media reports have suggested that the use of 3, 4-methylenedioxymethamphetamine (MDMA or 'ecstasy') is a significant problem across the United States. To date, however, available evidence has shown that the use of ecstasy has been concentrated among 'rave' attendees, with mainstream youth remaining relatively immune from its proliferation. In the current study we build on the ecstasy literature by examining the drug-using behaviors of 2,258 high school seniors with data collected through the Monitoring the Future program in 2002. Prevalence estimates of ecstasy use are generated and associations between ecstasy use, demographic characteristics, and alcohol and other drug use are explored. Ten percent of the sample reported lifetime ecstasy use, 7% reported use within the past 12 months, and 3% reported use within the past 30 days. Compared to non-users, 12-month ecstasy users were significantly more likely than non-users to be white and to have used all other drugs of abuse during the 12 months preceding the interview. These results suggest that ecstasy-related prevention efforts should target the at-risk population of rave attendees rather than the mainstream population of high school students. [ABSTRACT FROM AUTHOR]

This study offers cross-sectional analysis of data collected from the Alcohol and Substance Abuse Awareness and Use Study (ASAAUS). Data collection took place in October 2003 via a self-administered survey. Students who reported sexual abuse history in or around campus were 4 times more likely to report current amphetamine, cocaine, hallucinogen, opiate, or designer drug use compared to students with no sexual abuse history. Logistic regression analyses indicated that lifetime drug use was significantly associated with sexual abuse history (OR = 2.2, p < .05). While the relationships tested in this study are exploratory, they provide evidence for an important connection between sexual abuse and substance use among African-American college students. [ABSTRACT FROM AUTHOR]

The 'rave' phenomenon has been a major element in the resurgence of psychedelic drug use in Western society. Purportedly central to raves is the use of 'club drugs,' such as 3,4-methylenedioxymethamphetamine (MDMA or 'ecstasy'). To date, however, no studies have attempted to estimate MDMA dependence among rave attendees. In the current study, personal drug data were collected from 154 adult 'club rave' attendees along the Baltimore-Washington corridor between September and November 2002. The Center for Substance Abuse Research (CESAR) Arrestee Drug Screener (CADS) was used to screen for ecstasy dependence. Seventeen percent of the sample was diagnosed with probable MDMA-dependence. MDMA-dependent rave attendees were significantly more likely than non-dependents to be male, white, and to have used most illicit drugs during the 12 months preceding the interview. Logistic regression identified that gender, race, and past-year use of marijuana and PCP were the strongest predictors of MDMA dependence. Implications for these findings are discussed. [ABSTRACT FROM AUTHOR]

Anecdotal reports have suggested that the use and abuse of methamphetamine is increasing in the United States. To date, however, few scholarly works have explored the prevalence and correlates of methamphetamine use in the United States. To address this limitation, a secondary analysis was undertaken with data collected through the Arrestee Drug Abuse Monitoring (ADAM) Program. An examination of methamphetamine-positive rates among ADAM arrestees interviewed between 1991 and 2001 identified that its use is concentrated within the Western part of the United States. Supplementary analyses of Sacramento arrestees indicated that 71% were diagnosed as needing some form of alcohol or other drug treatment. Compared to those arrestees who were detected methamphetamine-negative, methamphetamine-positive arrestees in Sacramento were three times as likely to be white and to have been diagnosed in need of drug treatment. Implications for drug control policy are discussed. [ABSTRACT FROM AUTHOR]

This study explored job satisfaction and changes needed to help boost levels of job satisfaction. Self-reported job satisfaction data were collected from 71 school nurses employed in elementary, middle, and high school settings via interactive focus groups. The subjects participated in a 30- to 45-minute focus group session that was audiotaped and transcribed by the principal investigator. Beliefs about job satisfaction were identified and classified into exclusive categories or themes. While the majority of school nurses expressed contentment with their jobs, certain factors that would increase job satisfaction, such as salary and control issues, were discussed. Overall, 83% of school nurses in this study were satisfied in their present positions; however, issues of coping and role strain were identified as major contributors to low morale. Only 17% of the school nurses voiced job dissatisfaction, primarily attributed to low salaries and lack of trust and support from administration. As school nurses face a diverse community with complex needs, adaptation is needed for job satisfaction to be maintained. For this to occur, school nurses must take the initiative to educate administrators, parents, and communities about their role in the school setting. [ABSTRACT FROM AUTHOR]

A plethora of anecdotal evidence has suggested that use of 'club drugs,' such as lysergic acid diethylamide (LSD), Ketamine, Gamma hydroxybutyrate (GHB), and 3,4-methylenedioxy-methamphetamine (MDMA or 'ecstasy'), is a serious problem among youth across the United States. Unfortunately, little scientific evidence exists to support this contention. In the current study, we examine the ecstasy and other club drug (EOCD)-using behaviors of 1,561 high school seniors surveyed in 2000 through the Monitoring the Future (MTF) study. Prevalence estimates of EOCD use are generated and associations between EOCD use, demographic characteristics, and alcohol and other drug (AOD) use are explored. Twelve-moth self-reports for ecstasy, LSD, Ketamine, and GHB were 8%, 6%, 2%, and 1%, respectively. Compared to students who reported no use of EOCD during the 12 months preceding the interview, 12-month EOCD users were significantly more likely to be white, to live in the suburbs, and to have used all other drugs of abuse during the 12 months preceding the interview. Logistic regression identified that the strongest predictors of 12-month EOCD use were the use of heroin and other psychedelic drugs during the 12 months preceding the interview. Implications of these findings are discussed. [ABSTRACT FROM AUTHOR]

Self-report drug use data were collected from 282 female alternative school students surveyed through the Safer Choices 2 study in Houston, Texas. Data collection took place between October 2000 and March 2001 via audio-enabled laptop computers equipped with headphones. Logistic regression analyses indicated that sexual abuse history was significantly associated with lifetime use (OR = 1.9, p < or = 0.05). While the relationships tested in this study are exploratory, they provide evidence for an important connection between sexual abuse and substance use among female alternative school students. [ABSTRACT FROM AUTHOR]

A consistent 20-year old trend in adolescent tobacco use is that African American youth smoke cigarettes at a lower rate than other ethnic groups. To better understand this difference, our study identified gender and ethnic beliefs about cigarette smoking and abstention from smoking. In a multi-ethnic sample of adolescents. Based on qualitative analysis of 63 in depth interviews with African American, Hispanic, and European American adolescents, we found unique beliefs. In general, adolescents were well aware of adverse health effects of smoking. Female adolescents were more concerned with aesthetics as a smoking disadvantage than were male adolescents. Unique to this study, African Americans and Never Smokers emphasized moral and ethical reasons for not smoking. Examples of the Moral/Ethical theme are 'They know wrong from right' and 'It is not right to do it.' We discuss how the Moral/Ethical theme may have developed. Further, we present recommendations for future research to clarify influences for smoking and not smoking by gender and ethnic group among adolescents. [ABSTRACT FROM AUTHOR]

PURPOSE: Developing and disseminating innovative and effective approaches for smoking prevention among middle school children remains a public health priority. This pilot study evaluates the use of a Web-based tobacco prevention program, Headbutt, to change intentions of middle school children to smoke tobacco. METHODS: Headbutt was implemented with the use of a single-group pretest-posttest study design in sixth grade classes of nine middle schools in Texas (student n = 2227). The program assesses cognitive determinants of smoking and provides intervention feedback tailored to the child's responses. RESULTS: Headbutt significantly affected smoking intentions, prosmoking attitudes, self-efficacy expectations, and knowledge of negative consequences (all p < or = .001) measured with scales adopted from the Texas Tobacco Initiative Survey. Change in prosmoking attitudes had the greatest predictive effect on smoking intentions (p < .001). These results were moderated by ethnicity and age of students. CONCLUSION: Findings need to be interpreted in the light of study design limitations. However, strong associations between the Headbutt program and intention change suggests that a more rigorous effectiveness trial is indicated. [ABSTRACT FROM AUTHOR]

A growing body of recent research has identified that 'rave' attendees are at high risk for the use of 'club drugs,' such as 3,4-methylenedioxymethamphetamine (MDMA or 'ecstasy'). Rave attendees, however, comprise only one of several club-going populations. In the current study, we explore the prevalence of ecstasy and other club drug (EOCD) use among a sample of club attendees in Washington, DC. Data were collected from adult, primarily homosexual, club attendees during the summer of 2003. Data collection was scheduled between 11 p.m. and 3 a.m. Participation rates were high. Of the 211 club attendees approached, 88% (n = 186) completed the interview. Drug use prevalence rates were low. With the exception of alcohol and marijuana, 2-day self-reports were less than 1% for each drug. These findings, amalgamated with results from other EOCD-related studies involving several distinct populations, offer considerable insight into the state of ecstasy in American society. Based on a meta-analysis of this literature, we offer a community-level prevention intervention for the population at highest risk for EOCD use-rave attendees. [ABSTRACT FROM AUTHOR]

Land plants are well-known producers of terpenoids that play diverse roles in plant-environment interactions. The vast chemical diversity of terpenoids is initiated by terpene synthases. Plants contain a distinct mid-sized terpene synthase gene family termed TPS , which appears to have an ancient origin in a fused bacterial Class I (di)terpene synthase (TS) and Class II diterpene cyclase (DTC), corresponding to the catalytically relevant α-domain and βγ-didomains, respectively. However, while such fused tridomain bifunctional (Class I/II) diterpene cyclases/synthases (DCSs) have been found in plants (and fungi), no examples have been reported from bacteria, leaving the origin of the fusion event initiating the TPS gene family opaque. Here, the discovery of such tridomain bifunctional DCSs in bacteria is reported. Extensive genome mining unearthed five putative bacterial DCSs, with biochemical characterization revealing the expected bifunctional activity for three. The most intriguing was CseDCS from Candidatus sericytochromatia bacterium, which produces ent -kaurene, an intermediate in plant hormone biosynthesis, as this is the hypothesized activity for the ancestral TPS . Unlike the extant functionally equivalent TPSs , it was possible to split CseDCS into separate, independently acting DTC and TS, with the first producing the expected ent -copalyl diphosphate (CPP), serving as a CPP synthase (CPS), while the second converts this to ent -kaurene, serving as a kaurene synthase (KS). Nevertheless, sequence alignment and mutation analysis revealed intriguing similarities between this cyanobacterial fused CPS-KS and functionally equivalent TPSs . Regardless of the exact relationship, the discovery of fused bifunctional DCSs in bacteria supports the hypothesized origin of the plant TPS family from such a bacterial gene., Competing Interests: None declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nanjing Agricultural University.)

Rice ( Oryza sativa ) produces numerous diterpenoid phytoalexins that are important in defense against pathogens. Surprisingly, despite extensive previous investigations, a major group of such phytoalexins, the abietoryzins, were only recently reported. These aromatic abietanes are presumably derived from ent -miltiradiene, but such biosynthetic capacity has not yet been reported in O. sativa . While wild rice has been reported to contain such an enzyme, specifically ent -kaurene synthase-like 10 (KSL10), the only characterized ortholog from O. sativa ( OsKSL10 ), specifically from the well-studied cultivar (cv.) Nipponbare, instead has been shown to make ent -sandaracopimaradiene, precursor to the oryzalexins. Notably, in many other cultivars, OsKSL10 is accompanied by a tandem duplicate, termed here OsKSL14 . Biochemical characterization of OsKLS14 from cv. Kitaake demonstrates that this produces the expected abietoryzin precursor ent -miltiradiene. Strikingly, phylogenetic analysis of OsKSL10 across the rice pan-genome reveals that from cv. Nipponbare is an outlier, whereas the alleles from most other cultivars group with those from wild rice, suggesting that these also might produce ent -miltiradiene. Indeed, OsKSL10 from cv. Kitaake exhibits such activity as well, consistent with its production of abietoryzins but not oryzalexins. Similarly consistent with these results is the lack of abietoryzin production by cv. Nipponbare. Although their equivalent product outcome might suggest redundancy, OsKSL10 and OsKSL14 were observed to exhibit distinct expression patterns, indicating such differences may underlie retention of these duplicated genes. Regardless, the results reported here clarify abietoryzin biosynthesis and provide insight into the evolution of rice diterpenoid phytoalexins., Supplementary Information: The online version contains supplementary material available at 10.1007/s42994-024-00167-3., Competing Interests: Conflict of interestOn behalf of all the authors, the corresponding author states there are no conflicts of interest., (© The Author(s) 2024.)

Cytochromes P450 (CYPs) are heme-thiolate monooxygenases that prototypically catalyze the insertion of oxygen into unactivated C-H bonds but are capable of mediating more complex reactions. One of the most remarked-upon alternative reactions occurs during biosynthesis of the gibberellin A (GA) phytohormones, involving hydrocarbon ring contraction with coupled aldehyde extrusion of ent -kaurenoic acid to form the first gibberellin intermediate. While the unusual nature of this reaction has long been noted, its mechanistic basis has remained opaque. Building on identification of the relevant CYP114 from bacterial GA biosynthesis, detailed structure-function studies are reported here, including development of in vitro assays as well as crystallographic analyses both in the absence and presence of substrate. These structures provided insight into enzymatic catalysis of this unusual reaction, as exemplified by identification of a key role for the "missing" acid from an otherwise highly conserved acid-alcohol pair of residues. Notably, the results demonstrate that ring contraction requires dual factors, both the use of a dedicated ferredoxin and absence of the otherwise conserved acidic residue, with exclusion of either limiting turnover to just the initiating and more straightforward hydroxylation. The results provide detailed insight into the enzymatic structure-function relationships underlying this fascinating reaction and support the use of a semipinacol mechanism for the unusual ring contraction reaction.

Covering: up to 2014-2022.Diterpenoid biosynthesis in plants builds on the necessary production of ( E , E , E )-geranylgeranyl diphosphate (GGPP) for photosynthetic pigment production, with diterpenoid biosynthesis arising very early in land plant evolution, enabling stockpiling of the extensive arsenal of (di)terpenoid natural products currently observed in this kingdom. This review will build upon that previously published in the Annual Review of Plant Biology, with a stronger focus on enzyme structure-function relationships, as well as additional insights into the evolution of (di)terpenoid metabolism since generated.

Rice produces many diterpenoid phytoalexins and, reflecting the importance of these natural products in this important cereal crop plant, its genome contains three biosynthetic gene clusters ( BGCs ) for such metabolism. The chromosome 4 BGC ( c4BGC ) is largely associated with momilactone production, in part due to the presence of the initiating syn -copalyl diphosphate (CPP) synthase gene ( OsCPS4 ). Oryzalexin S is also derived from syn -CPP. However, the relevant subsequently acting syn -stemarene synthase gene ( OsKSL8 ) is not located in the c4BGC . Production of oryzalexin S further requires hydroxylation at carbons 2 and 19 (C2 and C19), presumably catalyzed by cytochrome P450 (CYP) monooxygenases. Here it is reported the closely related CYP99A2 and CYP99A3, whose genes are also found in the c4BGC catalyze the necessary C19-hydroxylation, while the closely related CYP71Z21 and CYP71Z22, whose genes are found in the recently reported chromosome 7 BGC ( c7BGC ), catalyze subsequent hydroxylation at C2α. Thus, oryzalexin S biosynthesis utilizes two distinct BGCs , in a pathway cross-stitched together by OsKSL8 . Notably, in contrast to the widely conserved c4BGC , the c7BGC is subspecies (ssp.) specific, being prevalent in ssp. japonica and only rarely found in the other major ssp. indica. Moreover, while the closely related syn -stemodene synthase OsKSL11 was originally considered to be distinct from OsKSL8 , it has now been reported to be a ssp. indica derived allele at the same genetic loci. Intriguingly, more detailed analysis indicates that OsKSL8(j) is being replaced by OsKSL11 ( OsKSL8i ), suggesting introgression from ssp. indica to (sub)tropical japonica, with concurrent disappearance of oryzalexin S production., Supplementary Information: The online version contains supplementary material available at 10.1007/s42994-022-00092-3., Competing Interests: Conflict of interestOn behalf of all authors, the corresponding author states that there is no conflict of interest., (© The Author(s) 2023.)

The devastating human pathogen Mycobacterium tuberculosis (Mtb) is able to parasitize phagosomal compartments within alveolar macrophage cells due, in part, to the activity of its cell-surface lipids. Prominent among these is 1-tuberculosinyl-adenosine (1-TbAd), a derivative of the diterpenoid tuberculosinyl (halima-5,13-dienyl) diphosphate produced by the class II diterpene cyclase encoded by Rv3377c, termed here MtHPS. Given the demonstrated ability of 1-TbAd to act as a virulence factor for Mtb and the necessity for Rv3377c for its production, there is significant interest in MtHPS activity. Class II diterpene cyclases catalyze a general acid-base-mediated carbocation cascade reaction initiated by protonation of the terminal alkene in the general diterpenoid precursor ( E,E,E )-geranylgeranyl diphosphate and terminated by deprotonation of the final cyclized (and sometimes also rearranged) intermediate. Here, structure-guided mutagenesis was applied to characterize the various residues contributing to activation of the enzymatic acid, as well as identify the enzymatic base in MtHPS. Particularly given the ability of conservative substitution for the enzymatic base (Y479F) to generate an alternative product (labda-7,13-dienyl diphosphate) via deprotonation of an earlier unrearranged intermediate, further mutational analysis was carried out to introduce potential alternative catalytic bases. The results were combined with mechanistic molecular modeling to elucidate how these mutations affect the catalytic activity of this important enzyme. This not only provided detailed structure-function insight into MtHPS but also further emphasized the inert nature of the active site of MtHPS and class II diterpene cyclases more generally., Competing Interests: The authors declare no competing financial interest., (© 2022 The Authors. Published by American Chemical Society.)

Rice diterpenoid phytoalexins (DPs) are secondary metabolites with a well known role in resistance to foliar pathogens. As DPs are also known to be produced and exuded by rice roots, we hypothesised that they might play an important role in plant-nematode interactions, and particularly in defence against phytoparasitic nematodes. We used transcriptome analysis on rice roots to analyse the effect of infection by the root-knot nematode Meloidogyne graminicola or treatment with resistance-inducing chemical stimuli on DP biosynthesis genes, and assessed the susceptibility of mutant rice lines impaired in DP biosynthesis to M. graminicola. Moreover, we grew these mutants and their wild-type in field soil and used metabarcoding to assess the effect of impairment in DP biosynthesis on rhizosphere and root nematode communities. We show that M. graminicola suppresses DP biosynthesis genes early in its invasion process and, conversely, that resistance-inducing stimuli transiently induce the biosynthesis of DPs. Moreover, we show that loss of DPs increases susceptibility to M. graminicola. Metabarcoding on wild-type and DP-deficient plants grown in field soil reveals that DPs significantly alter the composition of rhizosphere and root nematode communities. Diterpenoid phytoalexins are important players in basal and inducible defence against nematode pathogens of rice and help shape rice-associated nematode communities., (© 2022 The Authors. New Phytologist © 2022 New Phytologist Foundation.)

Plant-associated microbes have evolved the ability to independently produce gibberellin (GA) phytohormones as a mechanism to influence their host. Indeed, GA was first discovered as a metabolite from the fungal rice pathogen Gibberella fujikuroi, which uses it as a virulence factor. Though some bacterial plant pathogens similarly use GA to promote infection, symbiotic nitrogen-fixing bacteria (rhizobia), which inhabit the root nodules of legumes, also can produce GA, suggesting a role in symbiosis. The bacterial GA biosynthetic operon has been identified, but in rhizobia this typically no longer encodes the final metabolic gene (cyp115), so that these symbionts can only produce the penultimate intermediate GA 9 . Here, we demonstrate that soybean (Glycine max) expresses functional GA 3-oxidases (GA3ox) within its nodules, which have the capability to convert GA 9 produced by the enclosed rhizobial symbiont Bradyrhizobium diazoefficiens to bioactive GA 4 . This rhizobia-derived GA is demonstrated to cause an increase in nodule size and decrease in the number of nodules. The increase in individual nodule size correlates to greater numbers of bacterial progeny within a nodule, thereby providing a selective advantage to rhizobia that produce GA during the rhizobia-legume symbiosis. The expression of GA3ox in nodules and resultant nodulation effects of the GA product suggests that soybean has co-opted control of bioactive GA production, and thus nodule size, for its own benefit. Thus, our results suggest rhizobial GA biosynthesis has coevolved with host plant metabolism for cooperative production of a phytohormone that influences nodulation in a mutually beneficial manner., (© 2022. The Author(s), under exclusive licence to International Society for Microbial Ecology.)