Short prokaryotic Argonaute systems trigger cell death upon detection of invading DNA.

Argonaute proteins use single-stranded RNA or DNA guides to target complementary nucleic acids. This allows eukaryotic Argonaute proteins to mediate RNA interference and long prokaryotic Argonaute proteins to interfere with invading nucleic acids. The function and mechanisms of the phylogenetically distinct short prokaryotic Argonaute proteins remain poorly understood. We demonstrate that short prokaryotic Argonaute and the associated TIR-APAZ (SPARTA) proteins form heterodimeric complexes. Upon guide RNA-mediated target DNA binding, four SPARTA heterodimers form oligomers in which TIR domain-mediated NAD(P)ase activity is unleashed. When expressed in Escherichia coli, SPARTA is activated in the presence of highly transcribed multicopy plasmid DNA, which causes cell death through NAD(P)+ depletion. This results in the removal of plasmid-invaded cells from bacterial cultures. Furthermore, we show that SPARTA can be repurposed for the programmable detection of DNA sequences. In conclusion, our work identifies SPARTA as a prokaryotic immune system that reduces cell viability upon RNA-guided detection of invading DNA.

Thermus thermophilus Argonaute Functions in the Completion of DNA Replication.

In many eukaryotes, Argonaute proteins, guided by short RNA sequences, defend cells against transposons and viruses. In the eubacterium Thermus thermophilus, the DNA-guided Argonaute TtAgo defends against transformation by DNA plasmids. Here, we report that TtAgo also participates in DNA replication. In vivo, TtAgo binds 15- to 18-nt DNA guides derived from the chromosomal region where replication terminates and associates with proteins known to act in DNA replication. When gyrase, the sole T. thermophilus type II topoisomerase, is inhibited, TtAgo allows the bacterium to finish replicating its circular genome. In contrast, loss of gyrase and TtAgo activity slows growth and produces long sausage-like filaments in which the individual bacteria are linked by DNA. Finally, wild-type T. thermophilus outcompetes an otherwise isogenic strain lacking TtAgo. We propose that the primary role of TtAgo is to help T. thermophilus disentangle the catenated circular chromosomes generated by DNA replication.

High-throughput biochemical profiling reveals functional adaptation of a bacterial Argonaute.

Argonautes are nucleic acid-guided proteins that perform numerous cellular functions across all domains of life. Little is known about how distinct evolutionary pressures have shaped each Argonaute's biophysical properties. We applied high-throughput biochemistry to characterize how Thermus thermophilus Argonaute (TtAgo), a DNA-guided DNA endonuclease, finds, binds, and cleaves its targets. We found that TtAgo uses biophysical adaptations similar to those of eukaryotic Argonautes for rapid association but requires more extensive complementarity to achieve high-affinity target binding. Using these data, we constructed models for TtAgo association rates and equilibrium binding affinities that estimate the nucleic acid- and protein-mediated components of the target interaction energies. Finally, we showed that TtAgo cleavage rates vary widely based on the DNA guide, suggesting that only a subset of guides cleaves targets on physiologically relevant timescales.

Autonomous Generation and Loading of DNA Guides by Bacterial Argonaute.

Several prokaryotic Argonaute proteins (pAgos) utilize small DNA guides to mediate host defense by targeting invading DNA complementary to the DNA guide. It is unknown how these DNA guides are being generated and loaded onto pAgo. Here, we demonstrate that guide-free Argonaute from Thermus thermophilus (TtAgo) can degrade double-stranded DNA (dsDNA), thereby generating small dsDNA fragments that subsequently are loaded onto TtAgo. Combining single-molecule fluorescence, molecular dynamic simulations, and structural studies, we show that TtAgo loads dsDNA molecules with a preference toward a deoxyguanosine on the passenger strand at the position opposite to the 5' end of the guide strand. This explains why in vivo TtAgo is preferentially loaded with guides with a 5' end deoxycytidine. Our data demonstrate that TtAgo can independently generate and selectively load functional DNA guides.

Argonaute system of Kordia jejudonensis is a heterodimeric nucleic acid-guided nuclease.

We purified and characterized a prokaryotic argonaute (pAgo) (KjMP) and its associated protein (KjAA) from a bacterium Kordia jejudonensis. The two proteins present as a complex were revealed by the copurification of KjAA with His-tagged KjMP by Ni-NTA affinity column. The KjAA/KjMP complex was a heterodimer evaluated from the molecular weight estimated using size exclusion chromatography. The pAgo complex presented a guide-dependent target DNA cleavage. RNA was the preferred guide; however, DNA also functioned, albeit weakly. Additionally, 5'-phosphorylate or non-phosphorylated guide was equally effective. The purified complex exhibited nonspecific nuclease activity on dsDNA and ssDNA. This is the first study to report that short pAgo and its associated protein form a complex, which has a nucleic acid-guided target recognition and cleavage.

Recognition of double-stranded DNA by the Rhodobacter sphaeroides Argonaute protein.

In contrast to eukaryotic Argonaute proteins that act on RNA targets, prokaryotic Argonautes (pAgos) can target DNA, using either small RNA or small DNA guides for its recognition. Since pAgos can recognize only a single strand of DNA and lack a helicase activity, it remains unknown how double-stranded DNA can be bound both in vitro and in vivo. Here, using in vitro reconstitution and footprinting assays we analyze formation of specific complexes with target DNA by a catalytically inactive pAgo, RsAgo from Rhodobacter sphaeroides programmed with small guide RNAs. We showed that RsAgo can recognize a specific site in double-stranded DNA after stepwise reconstitution of the complex from individual oligonucleotides or after prior melting of the DNA target. When bound, RsAgo stabilizes an open DNA bubble corresponding to the length of the guide molecule and protects the target DNA from nuclease cleavage. The results suggest that RsAgo and, possibly, other RNA-guided pAgos cannot directly attack double-stranded DNA and likely require DNA opening by other cellular processes for their action.

Trends in biodiversity and habitat quantification tools used for market-based conservation in the United States.

Market-based conservation mechanisms are designed to facilitate the mitigation of harm to and conservation of habitats and biodiversity. Their potential is partly hindered, however, by the quantification tools used to assess habitat quality and functionality. Of specific concern are the lack of transparency and standardization in tool development and gaps in tool availability. To address these issues, we collected information via internet and literature searchers and through conversations with tool developers and users on tools used in U.S. conservation mechanisms, such as payments for ecosystem services (PES) and ecolabel programs, conservation banking, and habitat exchanges. We summarized information about tools and explored trends among and within mechanisms based on criteria detailing geographic, ecological, and technical features of tools. We identified 69 tools that assessed at least 34 species and 39 habitat types. Where tools reported pricing, 98% were freely available. More tools were applied to states along the U.S. West Coast than elsewhere, and the level of tool transferability varied markedly among mechanisms. Tools most often incorporated conditions at numerous spatial scales, frequently addressed multiple risks to site viability, and required 1-83 data inputs. Most tools required a moderate or greater level of user skill. Average tool-complexity estimates were similar among all mechanisms except PES programs. Our results illustrate the diversity among tools in their ecological features, data needs, and geographic application, supporting concerns about a lack of standardization. However, consistency among tools in user skill requirements, incorporation of multiple spatial scales, and complexity highlight important commonalities that could serve as a starting point for establishing more standardized tool development and feature-incorporation processes. Greater standardization in tool design may expand market participation and facilitate a needed assessment of the effectiveness of market-based conservation.

Tendencias en la Biodiversidad y en las Herramientas de Cuantificación de Hábitat Usadas para la Conservación Basada en el Mercado en los Estados Unidos Resumen Los mecanismos de conservación basada en el mercado están diseñados para facilitar la mitigación del daño y la conservación de los hábitats y la biodiversidad. Sin embargo, el potencial de estos mecanismos está parcialmente reducido por las herramientas de cuantificación usadas para evaluar la calidad y funcionalidad del hábitat. Son de preocupación específica la falta de transparencia y la estandarización del desarrollo de herramientas y los vacíos en la disponibilidad de las herramientas. Para tratar estos temas recolectamos información por medio del internet y los buscadores de literatura y a través de conversaciones con los desarrolladores y usuarios de las herramientas utilizadas en los mecanismos de conservación en los Estados Unidos, como la eco-etiqueta y los programas de pago por servicios ambientales (PES, en inglés), el banco de conservación y el intercambio de hábitats. Resumimos la información sobre las herramientas y exploramos tendencias entre y dentro de los mecanismos basados en criterios que detallan las características geográficas, ecológicas y técnicas de las herramientas. Identificamos 69 herramientas que evaluaron al menos a 34 especies y 39 tipos de hábitat. En donde las herramientas reportaron tarificación, el 98% estaban disponibles gratuitamente. Más herramientas estaban aplicadas a estados ubicados a lo largo de la costa oeste de los Estados Unidos que en cualquier otro sitio y el nivel de transferibilidad de las herramientas varió notablemente entre los mecanismos. Las herramientas comúnmente incorporaron las condiciones a numerosas escalas espaciales, trataron con frecuencia los múltiples riesgos para la viabilidad de sitio y requirieron 1 - 83 entradas de datos. La mayoría de las herramientas requirió un nivel moderado o mayor de habilidad para el usuario. Los estimados medios de la complejidad de las herramientas fueron similares entre todos los mecanismos, a excepción de los programas PES. Nuestros resultados ilustran la diversidad de características ecológicas, necesidades de datos y aplicación geográfica que existe entre las herramientas, lo que respalda las preocupaciones sobre la falta de estandarización. Sin embargo, la consistencia entre las herramientas en cuanto a los requerimientos de habilidades para el usuario, la incorporación de múltiples escalas espaciales y la complejidad resaltan las similitudes importantes que podrían servir como punto inicial para el establecimiento de un desarrollo más estandarizado de herramientas y procesos que incorporen las características del sitio. Una mayor estandarización del diseño de herramientas podría expandir la participación del mercado y facilitar una urgente evaluación de la efectividad de la conservación basada en el mercado.

Limits of monetization in protecting ecosystem services.

The monetary valuation of ecosystem services is gaining traction in policy and business communities. Several tools and decision-making processes have been proposed, including criteria to assess the appropriateness of using monetary valuation for biodiversity conservation outcomes. These criteria include measures such as scale, uniqueness, and threat. We used case studies of monetization projects for which the outcomes were measured to explore the limitations and application of these criteria. There was limited evidence of the effectiveness of such schemes. The majority of the schemes were established in areas where the criteria specifically excluded their use in isolation. Thus, although some aspects of monetization may be beneficial for biodiversity conservation, these schemes were not being used appropriately and require some quantitative minimum (or maximum) measurements to be applied through additional policy or governance measures to ensure biodiversity conservation outcomes.

Improving quality of care through payment for performance: examining effects on the availability and stock-out of essential medical commodities in Tanzania.

OBJECTIVE: To evaluate the effects of payment for performance (P4P) on the availability and stock-out rate of reproductive, maternal, newborn and child health (RMNCH) medical commodities in Tanzania and assess the distributional effects. METHODS: The availability of RMNCH commodities (medicines, supplies and equipment) on the day of the survey, and stock-outs for at least one day in the 90 days prior to the survey, was measured in 75 intervention and 75 comparison facilities in January 2012 and 13 months later. Composite scores for each subgroup of commodities were generated. A difference-in-differences linear regression was used to estimate the effect of P4P on outcomes and differential effects by facility location, level of care, ownership and socio-economic status of the catchment population. RESULTS: We estimated a significant increase in the availability of medicines by 8.4 percentage points (P = 0.002) and an 8.3 percentage point increase (P = 0.050) in the availability of medical supplies. P4P had no effect on the availability of functioning equipment. Most items with a significant increase in availability also showed a significant reduction in stock-outs. Effects were generally equally distributed across facilities, with effects on stock-outs of many medicines being pro-poor, and greater effects in facilities in rural compared to urban districts. CONCLUSION: P4P can improve the availability of medicines and medical supplies, especially in poor, rural areas, when these commodities are incentivised at both facility and district levels, making services more acceptable, effective and affordable, enhancing progress towards universal health coverage.

Generating spatially optimized habitat in a trade-off between social optimality and budget efficiency.

Auctions have been proposed as alternatives to payments for environmental services when spatial interactions and costs are better known to landowners than to the conservation agency (asymmetric information). Recently, an auction scheme was proposed that delivers optimal conservation in the sense that social welfare is maximized. I examined the social welfare and the budget efficiency delivered by this scheme, where social welfare represents the difference between the monetized ecological benefit and the conservation cost incurred to the landowners and budget efficiency is defined as maximizing the ecological benefit for a given conservation budget. For the analysis, I considered a stylized landscape with land patches that can be used for agriculture or conservation. The ecological benefit was measured by an objective function that increases with increasing number and spatial aggregation of conserved land patches. I compared the social welfare and the budget efficiency of the auction scheme with an agglomeration payment, a policy scheme that considers spatial interactions and that was proposed recently. The auction delivered a higher level of social welfare than the agglomeration payment. However, the agglomeration payment was more efficient budgetarily than the auction, so the comparative performances of the 2 schemes depended on the chosen policy criterion-social welfare or budget efficiency. Both policy criteria are relevant for conservation. Which one should be chosen depends on the problem at hand, for example, whether social preferences should be taken into account in the decision of how much money to invest in conservation or whether the available conservation budget is strictly limited.

Impacts of a new insurance benefit with capitated provider payment on healthcare utilization, expenditure and quality of medication prescribing in China.

OBJECTIVES: To assess a new Chinese insurance benefit with capitated provider payment for common diseases in outpatients. METHODS: Longitudinal health insurance claims data, health administrative data and primary care facility data were used to assess trajectories in outpatient visits, inpatient admissions, expenditure per common disease outpatient (CD/OP) visit and prescribing indicators over time. We conducted segmented regression analyses of interrupted time series data to measure changes in level and trend overtime, and cross-sectional comparisons against external standards. RESULTS: The number of total outpatient visits at 46 primary care facilities (on the CD/OP benefit as of July 2012) increased by 46 895 visits/month (P = 0.004, 95% CI: 15 795-77 994); the average number of CD/OP visits reached 1.84/year/enrollee in 2012; monthly inpatient admissions dropped from 6.4 (2009) to 4.3 (2012) per 1000 enrollees; the median total expenditure per CD/OP visit dropped by CNY 15.40 (P = 0.16, 95% CI: -36.95~6.15); injectable use dropped by 7.38% (P = 0.03, 95% CI: -14.08%~-0.68%); antibiotic use was not improved. CONCLUSIONS: Zhuhai's new CD/OP benefit with capitated provider payment has expanded access to primary care, which may have led to a reduction in expensive specialist inpatient services for CD/OP benefit enrollees. Cost awareness was likely raised, and rapidly growing expenditures were contained. Although having been partially improved, inappropriate prescribing of antibiotics and injectables was still prevalent. More explicit incentives and specific quality of care targets must be incorporated into the capitated provider payment to promote scientifically sound and cost-effective care and treatment.

Cost-effectiveness of conservation payment schemes for species with different range sizes.

Payments to compensate landowners for carrying out costly land-use measures that benefit endangered biodiversity have become an important policy instrument. When designing such payments, it is important to take into account that spatially connected habitats are more valuable for many species than isolated ones. One way to incentivize provision of connected habitats is to offer landowners an agglomeration bonus, that is, a bonus on top of payments they are receiving to conserve land if the land is spatially connected. Researchers have compared the cost-effectiveness of the agglomeration bonus with 2 alternatives: an all-or-nothing, agglomeration payment, where landowners receive a payment only if the conserved land parcels have a certain level of spatial connectivity, and a spatially homogeneous payment, where landowners receive a payment for conserved land parcels irrespective of their location. Their results show the agglomeration bonus is rarely the most cost-effective option, and when it is, it is only slightly better than one of the alternatives. This suggests that the agglomeration bonus should not be given priority as a policy design option. However, this finding is based on consideration of only 1 species. We examined whether the same applied to 2 species, one for which the homogeneous payment is best and the other for which the agglomeration payment is most cost-effective. We modified a published conceptual model so that we were able to assess the cost-effectiveness of payment schemes for 2 species and applied it to a grassland bird and a grassland butterfly in Germany that require the same habitat but have different spatial-connectivity needs. When conserving both species, the agglomeration bonus was more cost-effective than the agglomeration and the homogeneous payment; thus, we showed that as a policy the agglomeration bonus is a useful conservation-payment option.

Five challenges to reconcile agricultural land use and forest ecosystem services in Southeast Asia.

Southeast Asia possesses the highest rates of tropical deforestation globally and exceptional levels of species richness and endemism. Many countries in the region are also recognized for their food insecurity and poverty, making the reconciliation of agricultural production and forest conservation a particular priority. This reconciliation requires recognition of the trade-offs between competing land-use values and the subsequent incorporation of this information into policy making. To date, such reconciliation has been relatively unsuccessful across much of Southeast Asia. We propose an ecosystem services (ES) value-internalization framework that identifies the key challenges to such reconciliation. These challenges include lack of accessible ES valuation techniques; limited knowledge of the links between forests, food security, and human well-being; weak demand and political will for the integration of ES in economic activities and environmental regulation; a disconnect between decision makers and ES valuation; and lack of transparent discussion platforms where stakeholders can work toward consensus on negotiated land-use management decisions. Key research priorities to overcome these challenges are developing easy-to-use ES valuation techniques; quantifying links between forests and well-being that go beyond economic values; understanding factors that prevent the incorporation of ES into markets, regulations, and environmental certification schemes; understanding how to integrate ES valuation into policy making processes, and determining how to reduce corruption and power plays in land-use planning processes.

Policy challenges and approaches for the conservation of mangrove forests in Southeast Asia.

Many drivers of mangrove forest loss operate over large scales and are most effectively addressed by policy interventions. However, conflicting or unclear policy objectives exist at multiple tiers of government, resulting in contradictory management decisions. To address this, we considered four approaches that are being used increasingly or could be deployed in Southeast Asia to ensure sustainable livelihoods and biodiversity conservation. First, a stronger incorporation of mangroves into marine protected areas (that currently focus largely on reefs and fisheries) could resolve some policy conflicts and ensure that mangroves do not fall through a policy gap. Second, examples of community and government comanagement exist, but achieving comanagement at scale will be important in reconciling stakeholders and addressing conflicting policy objectives. Third, private-sector initiatives could protect mangroves through existing and novel mechanisms in degraded areas and areas under future threat. Finally, payments for ecosystem services (PES) hold great promise for mangrove conservation, with carbon PES schemes (known as blue carbon) attracting attention. Although barriers remain to the implementation of PES, the potential to implement them at multiple scales exists. Closing the gap between mangrove conservation policies and action is crucial to the improved protection and management of this imperiled coastal ecosystem and to the livelihoods that depend on them.

Complementary benefits of tourism and hunting to communal conservancies in Namibia.

Tourism and hunting both generate substantial revenues for communities and private operators in Africa, but few studies have quantitatively examined the trade-offs and synergies that may result from these two activities. We evaluated financial and in-kind benefit streams from tourism and hunting on 77 communal conservancies in Namibia from 1998 to 2013, where community-based wildlife conservation has been promoted as a land-use that complements traditional subsistence agriculture. We used data collected annually for all communal conservancies to characterize whether benefits were derived from hunting or tourism. We classified these benefits into 3 broad classes and examined how benefits flowed to stakeholders within communities under the status quo and under a simulated ban on hunting. Across all conservancies, total benefits from hunting and tourism increased at roughly the same rate, although conservancies typically started generating benefits from hunting within 3 years of formation as opposed to after 6 years for tourism. Disaggregation of data revealed that the main benefits from hunting were income for conservancy management and food in the form of meat for the community at large. The majority of tourism benefits were salaried jobs at lodges. A simulated ban on trophy hunting significantly reduced the number of conservancies that could cover their operating costs, whereas eliminating income from tourism did not have as severe an effect. Given that the benefits generated from hunting and tourism typically begin at different times in a conservancy's life-span (earlier vs. later, respectively) and flow to different segments of local communities, these 2 activities together may provide the greatest incentives for conservation on communal lands in Namibia. A singular focus on either hunting or tourism would reduce the value of wildlife as a competitive land-use option and have grave repercussions for the viability of community-based conservation efforts in Namibia, and possibly other parts of Africa.

Trends in deforestation and forest degradation after a decade of monitoring in the Monarch Butterfly Biosphere Reserve in Mexico.

We used aerial photographs, satellite images, and field surveys to monitor forest cover in the core zones of the Monarch Butterfly Biosphere Reserve in Mexico from 2001 to 2012. We used our data to assess the effectiveness of conservation actions that involved local, state, and federal authorities and community members (e.g., local landowners and private and civil organizations) in one of the world's most iconic protected areas. From 2001 through 2012, 1254 ha were deforested (i.e., cleared areas had <10% canopy cover), 925 ha were degraded (i.e., areas for which canopy forest decreased), and 122 ha were affected by climatic conditions. Of the total 2179 ha of affected area, 2057 ha were affected by illegal logging: 1503 ha by large-scale logging and 554 ha by small-scale logging. Mexican authorities effectively enforced efforts to protect the monarch reserve, particularly from 2007 to 2012. Those efforts, together with the decade-long financial support from Mexican and international philanthropists and businesses to create local alternative-income generation and employment, resulted in the decrease of large-scale illegal logging from 731 ha affected in 2005-2007 to none affected in 2012, although small-scale logging is of growing concern. However, dire regional social and economic problems remain, and they must be addressed to ensure the reserve's long-term conservation. The monarch butterfly (Danaus plexippus) overwintering colonies in Mexico-which engage in one of the longest known insect migrations-are threatened by deforestation, and a multistakeholder, regional, sustainable-development strategy is needed to protect the reserve.

Do family physicians need more payment for working better? Financial incentives in primary care.

INTRODUCTION: Financial incentives are widely used in health services to improve the quality of care or to reach some specific targets. Pay for performance systems were also introduced in the primary health care systems of many European countries. OBJECTIVE: Our study aims to describe and compare recent existing primary care indicators and related financing in European countries. METHODS: Literature search was performed and questionnaires were sent to primary care experts of different countries within the European General Practice Research Network. RESULTS: Ten countries have published primary care quality indicators (QI) associated with financial incentives. The number of QI varies from 1 to 134 and can modify the finances of physicians with up to 25% of their total income. CONCLUSIONS: The implementations of these schemes should be critically evaluated with continuous monitoring at national or regional level; comparison is required between targets and their achievements, health gains and use of resources as well.

Unusual Guide-binding Pockets in RNA-targeting pAgo Nucleases.

Argonaute nucleases use small nucleic acid guides to recognize and degrade complementary nucleic acid targets. Most prokaryotic Argonautes (pAgos) recognize DNA targets and may play a role in cell immunity against invader genetic elements. We have recently described two related groups of pAgo nucleases that have distinct specificity for DNA guides and RNA targets (DNA > RNA pAgos). Here, we describe additional pAgos from the same clades of the pAgo tree and demonstrate that they have the same unusual nucleic acid specificity. The two groups of DNA > RNA pAgos have non-standard guide-binding pockets in the MID domain and differ in the register of guide DNA binding and target cleavage. In contrast to other pAgos, which coordinate the 5'-end of the guide molecule by their C-terminal carboxyl, DNA > RNA pAgos have an extended C-terminus located away from the MID pocket. We show that modifications of the C-terminus do not affect guide DNA binding, but inhibit cleavage of complementary and mismatched RNA targets by some DNA > RNA pAgos. Our data suggest that the unique C-terminus found in DNA > RNA pAgos can modulate their catalytic properties and can be used as a target for pAgo modifications.

Effect on hospital incentive payments and quality performance of a hospital pay for performance (P4P) programme in Belgium.

BACKGROUND: Belgium initiated a hospital pay for performance (P4P) programme after a decade of fixed bonus budgets for "quality and safety contracts". This study examined the effect of P4P on hospital incentive payments, performance on quality measures, and the association between changes in quality performance and incentive payments over time. METHODS: The Belgian government provided information on fixed bonus budgets in 2013-2017 and hospital incentive payments as well as hospital performance on quality measures for the P4P programmes in 2018-2020. Descriptive analyses were conducted to map the financial repercussion between the two systems. A difference-in-difference analysis evaluated the association between quality indicator performance and received incentive payments over time. RESULTS: Data from 87 acute-care hospitals were analyzed. In the transition to a P4P programme, 29% of hospitals received lower incentive payments per bed. During the P4P years, quality performance scores increased yearly for 55% of hospitals and decreased yearly for 5% of hospitals. There was a significant larger drop in incentive payments for hospitals that scored above median with the start of the P4P programme. CONCLUSIONS: The transition from fixed bonus budgets for quality efforts to a new incentive payment in a P4P programme has led to more hospitals being financially impacted, although the effect is marginal given the small P4P budget. Quality indicators seem to improve over the years, but this does not correlate with an increase in reward per bed for all hospitals due to the closed nature of the budget.

A long look at short prokaryotic Argonautes.

Argonaute proteins (Agos) use small 15-30 nucleotide-long guides to bind and/or cleave complementary target nucleic acids. Eukaryotic Agos mediate RNA-guided RNA silencing, while 'long' prokaryotic Agos (pAgos) use RNA or DNA guides to interfere with invading plasmid and viral DNA. Here, we review the function and mechanisms of truncated and highly divergent 'short' pAgos, which, until recently, remained functionally uncharacterized. Short pAgos have retained the Middle (MID) and P-element-Induced Wimpy Testis (PIWI) domains important for guide-mediated target binding, but lack the ability to cleave their targets. Instead, emerging insights reveal that various short pAgos interact with distinct accessory 'effector' enzymes. Upon guide-mediated detection of invading DNA by short pAgos, their associated effector enzymes kill the host cell and, consequentially, prevent spread of the invader.