Flagellar energy costs across the tree of life.

Flagellar-driven motility grants unicellular organisms the ability to gather more food and avoid predators, but the energetic costs of construction and operation of flagella are considerable. Paths of flagellar evolution depend on the deviations between fitness gains and energy costs. Using structural data available for all three major flagellar types (bacterial, archaeal, and eukaryotic), flagellar construction costs were determined for Escherichia coli, Pyrococcus furiosus, and Chlamydomonas reinhardtii. Estimates of cell volumes, flagella numbers, and flagellum lengths from the literature yield flagellar costs for another ~200 species. The benefits of flagellar investment were analysed in terms of swimming speed, nutrient collection, and growth rate; showing, among other things, that the cost-effectiveness of bacterial and eukaryotic flagella follows a common trend. However, a comparison of whole-cell costs and flagellum costs across the Tree of Life reveals that only cells with larger cell volumes than the typical bacterium could evolve the more expensive eukaryotic flagellum. These findings provide insight into the unsolved evolutionary question of why the three domains of life each carry their own type of flagellum.

Most creatures on Earth are single cell organisms. The tree of life comprises three domains, two of which ­ bacteria and archaea ­ are formed exclusively of creatures that spend their existence as independent cells. Yet even eukaryotes, the domain which include animals and plants, feature single cell species such as yeasts and algae. Regardless of which group they belong to, all single-celled organisms must find food in their environment. For this, many are equipped with flagella, whip-like structures that protrude from the cell and allow it to swim. In fact, archaea, bacteria and eukaryotes have all independently evolved these structures. However, flagella are also expensive for an organism to build, maintain and operate. They are only worth having if the advantages they bring to the cell compensate for their cost; many single-cell species do not carry flagella and obtain their food without having to swim. To explore this trade-off, Schavemaker and Lynch calculated the cost of building and using flagella for about 200 species across the tree of life. The analysis show that the amount of energy spent on flagella varied between 0.1% and 40% of the entire cell budget. This investment is only worthwhile if the cell is above a certain size. Smaller than this, and the organism is better off obtaining its food passively. The results also show that while eukaryotic flagella are much bigger and quite different than their bacterial counterpart, both appendages share the same patterns of cost effectiveness. However only eukaryotic cells, which are on average larger than bacteria, can afford to evolve such sizable and complex structures; making just one would cost more than the entire energy budget of a bacterial cell. Many single-cell species which are critical for the health of the planet are equipped with flagella, such as the microorganisms which recycle matter in the oceans and release carbon dioxide. Understanding the costs and benefits of flagella could explain more about this aspect of the carbon cycle, and therefore global warming.

Improved bounds on entropy production in living systems.

Living systems maintain or increase local order by working against the second law of thermodynamics. Thermodynamic consistency is restored as they consume free energy, thereby increasing the net entropy of their environment. Recently introduced estimators for the entropy production rate have provided major insights into the efficiency of important cellular processes. In experiments, however, many degrees of freedom typically remain hidden to the observer, and, in these cases, existing methods are not optimal. Here, by reformulating the problem within an optimization framework, we are able to infer improved bounds on the rate of entropy production from partial measurements of biological systems. Our approach yields provably optimal estimates given certain measurable transition statistics. In contrast to prevailing methods, the improved estimator reveals nonzero entropy production rates even when nonequilibrium processes appear time symmetric and therefore may pretend to obey detailed balance. We demonstrate the broad applicability of this framework by providing improved bounds on the energy consumption rates in a diverse range of biological systems including bacterial flagella motors, growing microtubules, and calcium oscillations within human embryonic kidney cells.

Long-term segmentation-free assessment of head-flagellum movement and intracellular calcium in swimming human sperm.

Human spermatozoa are the archetype of long-term self-organizing transport in nature and are critical for reproductive success. They utilize coordinated head and flagellar movements to swim long distances within the female reproductive tract in order to find and fertilize the egg. However, to date, long-term analysis of the sperm head-flagellar movements, or indeed those of other flagellated microorganisms, remains elusive due to limitations in microscopy and flagellar-tracking techniques. Here, we present a novel methodology based on local orientation and isotropy of bio-images to obtain long-term kinematic and physiological parameters of individual free-swimming spermatozoa without requiring image segmentation (thresholding). This computer-assisted segmentation-free method evaluates, for the first time, characteristics of the head movement and flagellar beating for up to 9.2 min. We demonstrate its powerful use by showing how releasing Ca2+ from internal stores significantly alters long-term sperm behavior. The method allows for straightforward generalization to other bio-imaging applications, such as studies of bull sperm and Trypanosoma, or indeed of other flagellated microorganisms - appealing to communities other than those investigating sperm biology.

A Screen for Antibiotic Resistance Determinants Reveals a Fitness Cost of the Flagellum in Pseudomonas aeruginosa.

The intrinsic resistance of Pseudomonas aeruginosa to many antibiotics limits treatment options for pseudomonal infections. P. aeruginosa's outer membrane is highly impermeable and decreases antibiotic entry into the cell. We used an unbiased high-throughput approach to examine mechanisms underlying outer membrane-mediated antibiotic exclusion. Insertion sequencing (INSeq) identified genes that altered fitness in the presence of linezolid, rifampin, and vancomycin, antibiotics to which P. aeruginosa is intrinsically resistant. We reasoned that resistance to at least one of these antibiotics would depend on outer membrane barrier function, as previously demonstrated in Escherichia coli and Vibrio cholerae This approach demonstrated a critical role of the outer membrane barrier in vancomycin fitness, while efflux pumps were primary contributors to fitness in the presence of linezolid and rifampin. Disruption of flagellar assembly or function was sufficient to confer a fitness advantage to bacteria exposed to vancomycin. These findings clearly show that loss of flagellar function alone can confer a fitness advantage in the presence of an antibiotic.IMPORTANCE The cell envelopes of Gram-negative bacteria render them intrinsically resistant to many classes of antibiotics. We used insertion sequencing to identify genes whose disruption altered the fitness of a highly antibiotic-resistant pathogen, Pseudomonas aeruginosa, in the presence of antibiotics usually excluded by the cell envelope. This screen identified gene products involved in outer membrane biogenesis and homeostasis, respiration, and efflux as important contributors to fitness. An unanticipated fitness cost of flagellar assembly and function in the presence of the glycopeptide antibiotic vancomycin was further characterized. These findings have clinical relevance for individuals with cystic fibrosis who are infected with P. aeruginosa and undergo treatment with vancomycin for a concurrent Staphylococcus aureus infection.

Acoustic trap-and-release for rapid assessment of cell motility.

Functional cilia and flagella are crucial to the propulsion of physiological fluids, motile cells, and microorganisms. Motility assessment of individual cells allows discrimination of normal from dysfunctional behavior, but cell-scale analysis of individual trajectories to represent a population is laborious and impractical for clinical, industrial, and even research applications. We introduce an assay that quantifies swimming capability as a function of the variation in polar moment of inertia of cells released from an acoustic trap. Acoustic confinement eliminates the need to trace discrete trajectories and enables automated analysis of hundreds of cells in minutes. The approach closely approximates the average speed estimated from the mean squared displacement of individual cells for wild-type Chlamydomonas reinhardtii and two mutants (ida3 and oda5) that display aberrant swimming behaviors. Large-population acoustic trap-and-release rapidly differentiates these cell types based on intrinsic motility, which provides a highly sensitive and efficient alternative to conventional particle tracing.

Stochastic Dynamics of Eukaryotic Flagellar Growth.

We study the dynamics of flagellar growth in eukaryotes where intraflagellar transporters (IFT) play a crucial role. First we investigate a stochastic version of the original balance point model where a constant number of IFT particles move up and down the flagellum. The detailed model is a discrete event vector-valued Markov process occurring in continuous time. First the detailed stochastic model is compared and contrasted with a simple scalar ordinary differential equation (ODE) model of flagellar growth. Numerical simulations reveal that the steady-state mean value of the stochastic model is well approximated by the ODE model. Then we derive a scalar stochastic differential equation (SDE) as a first approximation and obtain a "small noise" approximation showing flagellar length to be Gaussian with mean and variance governed by simple ODEs. The accuracy of the small noise model is compared favorably with the numerical simulation results of the detailed model. Secondly, we derive a revised SDE for flagellar length following the revised balance point model proposed in 2009 in which IFT particles move in trains instead of in isolation. Small noise approximation of the revised SDE yields the same approximate Gaussian distribution for the flagellar length as the SDE corresponding to the original balance point model.

Assessment of the flagellar redox potential in Chlamydomonas reinhardtii using a redox-sensitive fluorescent protein, Oba-Qc.

Cellular reducing-oxidizing (redox) potential is mainly determined by the concentration ratio between reduced and oxidized glutathiones. It is normally kept at a moderately reduced state but affected to some extent by metabolic activities such as respiration and/or photosynthesis. Changes in redox potential induce many cellular activities collectively called redox responses. For an understanding of the dynamics of the cellular redox responses, redox potential must be accurately assessed in vivo. In this study, we developed a method to measure the in vivo redox potential in the green alga Chlamydomonas reinhardtii, using Oba-Qc, a recently developed redox-monitoring protein. Taking advantage of the periodic flagellar assembly, we introduced Oba-Qc molecules into the flagella at a constant density. Fluorescence signals from flagella in live cells, calibrated against the fluorescence from the samples in buffers of known redox potentials, determined the redox potential to be ∼-250 mV in the light and ∼-280 mV in the dark. Introduction of a sensor protein fused with a structural protein that assembles at a constant density will be also applicable for measurements of various kinds cellular signals in flagella.

Single-molecule tracking in live Yersinia enterocolitica reveals distinct cytosolic complexes of injectisome subunits.

In bacterial type 3 secretion, substrate proteins are actively transported from the bacterial cytoplasm into the host cell cytoplasm by a large membrane-embedded machinery called the injectisome. Injectisomes transport secretion substrates in response to specific environmental signals, but the molecular details by which the cytosolic secretion substrates are selected and transported through the type 3 secretion pathway remain unclear. Secretion activity and substrate selectivity are thought to be controlled by a sorting platform consisting of the proteins SctK, SctQ, SctL, and SctN, which together localize to the cytoplasmic side of membrane-embedded injectisomes. However, recent work revealed that sorting platform proteins additionally exhibit substantial cytosolic populations and that SctQ reversibly binds to and dissociates from the cytoplasmic side of membrane-embedded injectisomes. Based on these observations, we hypothesized that dynamic molecular turnover at the injectisome and cytosolic assembly among sorting platform proteins is a critical regulatory component of type 3 secretion. To determine whether sorting platform complexes exist in the cytosol, we measured the diffusive properties of the two central sorting platform proteins, SctQ and SctL, using live cell high-throughput 3D single-molecule tracking microscopy. Single-molecule trajectories, measured in wild-type and mutant Yersinia enterocolitica cells, reveal that both SctQ and SctL exist in several distinct diffusive states in the cytosol, indicating that these proteins form stable homo- and hetero-oligomeric complexes in their native environment. Our findings provide the first diffusive state-resolved insights into the dynamic regulatory network that interfaces stationary membrane-embedded injectisomes with the soluble cytosolic components of the type 3 secretion system.

Preparation of gold nanoparticles supported on graphene oxide with flagella as the template for nonenzymatic hydrogen peroxide sensing.

Gold nanoparticles supported on graphene oxide with flagella as the template were developed as an electrochemical sensor for the detection of hydrogen peroxide (H2O2) in serum. The flagella-Au nanoparticles composite and graphene oxide were dropped onto a glassy carbon electrode (GCE) to form a new H2O2 electrochemical sensor. The structure morphology of the prepared sensor was characterized by transmission electron microscopy (TEM), and the electrocatalytic performance towards H2O2 reduction was evaluated by cyclic voltammetry (CV) and amperometric methods. The response current of the sensor showed a good linear relationship with the concentration of H2O2 in the range of 10-1000 µM (R2 = 0.9916). The minimum detection limit of 1 µM was obtained (S/N = 3). Finally, the sensor was applied to the detection of H2O2 in serum, and the recoveries were satisfactory. As the sensor is sensitive, fast, and easy to make, it is expected to be used for rapid detection of H2O2. Graphical abstract ᅟ.

Parasite motility is critical for virulence of African trypanosomes.

African trypanosomes, Trypanosoma brucei spp., are lethal pathogens that cause substantial human suffering and limit economic development in some of the world's most impoverished regions. The name Trypanosoma ("auger cell") derives from the parasite's distinctive motility, which is driven by a single flagellum. However, despite decades of study, a requirement for trypanosome motility in mammalian host infection has not been established. LC1 is a conserved dynein subunit required for flagellar motility. Prior studies with a conditional RNAi-based LC1 mutant, RNAi-K/R, revealed that parasites with defective motility could infect mice. However, RNAi-K/R retained residual expression of wild-type LC1 and residual motility, thus precluding definitive interpretation. To overcome these limitations, here we generate constitutive mutants in which both LC1 alleles are replaced with mutant versions. These double knock-in mutants show reduced motility compared to RNAi-K/R and are viable in culture, but are unable to maintain bloodstream infection in mice. The virulence defect is independent of infection route but dependent on an intact host immune system. By comparing different mutants, we also reveal a critical dependence on the LC1 N-terminus for motility and virulence. Our findings demonstrate that trypanosome motility is critical for establishment and maintenance of bloodstream infection, implicating dynein-dependent flagellar motility as a potential drug target.

Structure and Intermolecular Interactions between L-Type Straight Flagellar Filaments.

Bacterial mobility is powered by rotation of helical flagellar filaments driven by rotary motors. Flagellin isolated from the Salmonella Typhimurium SJW1660 strain, which differs by a point mutation from the wild-type strain, assembles into straight filaments in which flagellin monomers are arranged in a left-handed helix. Using small-angle x-ray scattering and osmotic stress methods, we investigated the structure of SJW1660 flagellar filaments as well as the intermolecular forces that govern their assembly into dense hexagonal bundles. The scattering data were fitted to models, which took into account the atomic structure of the flagellin subunits. The analysis revealed the exact helical arrangement and the super-helical twist of the flagellin subunits within the filaments. Under osmotic stress, the filaments formed two-dimensional hexagonal bundles. Monte Carlo simulations and continuum theories were used to analyze the scattering data from hexagonal arrays, revealing how the bundle bulk modulus and the deflection length of filaments in the bundles depend on the applied osmotic stress. Scattering data from aligned flagellar bundles confirmed the theoretically predicated structure-factor scattering peak line shape. Quantitative analysis of the measured equation of state of the bundles revealed the contributions of electrostatic, hydration, and elastic interactions to the intermolecular forces associated with bundling of straight semi-flexible flagellar filaments.

Production, characterization, and assessment of a stable analog of the response regulator CheY-phosphate from Thermotoga maritima.

Phosphorylation of CheY promotes association with the flagellar motor and ultimately controls the directional bias of the motor. However, biochemical studies of activated CheY-phosphate have been challenging due to the rapid hydrolysis of the aspartyl-phosphate in vitro. An inert analog of Tm CheY-phosphate, phosphono-CheY, was synthesized by chemical modification and purified by cation-exchange chromatography. Changes in HPLC retention times, chemical assays for phosphate and free thiol, and mass spectrometry experiments demonstrate modification of Cys54 with a phosphonomethyl group. Additionally, a crystal structure showed electron density for the phosphonomethyl group at Cys54, consistent with a modification at that position. Subsequent biochemical experiments confirmed that protein crystals were phosphono-CheY. Isothermal titration calorimetry and fluorescence polarization binding assays demonstrated that phosphono-CheY bound a peptide derived from FliM, a native partner of CheY-phosphate, with a dissociation constant of ∼29 µM, at least sixfold more tightly than unmodified CheY. Taken together these results suggest that Tm phosphono-CheY is a useful and unique analog of Tm CheY-phosphate.

Energy cost and putative benefits of cellular mechanisms modulating buoyancy in aflagellate marine phytoplankton.

Little information is available on the energetics of buoyancy modulation in aflagellate phytoplankton, which comprises the majority of autotrophic cells found in the ocean. Here, we computed for three aflagellate species of marine phytoplankton (Emiliania huxleyi, Thalassiosira pseudonana, and Ethmodiscus rex) the theoretical minimum energy cost as photons absorbed and nitrogen resource required of the key physiological mechanisms (i.e., replacement of quaternary ammonium by dimethyl-sulfoniopropionate, storage of polysaccharides, and cell wall biosynthesis) affecting the cell's vertical movement as a function of nitrogen (N) availability. These energy costs were also normalized to the capacity of each buoyancy mechanism to modulate sinking or rising rates based on Stokes' law. The three physiological mechanisms could act as ballast in the three species tested in conditions of low N availability at a low fraction (<12%) of the total photon energy cost for growth. Cell wall formation in E. huxleyi was the least costly ballast strategy, whereas in T. pseudonana, the photon energy cost of the three ballast strategies was similar. In E. rex, carbohydrate storage and mobilization appear to be energetically cheaper than modulations in organic solute synthesis to achieve vertical migration. This supports the carbohydrate-ballast strategy for vertical migration for this species, but argues against the theory of replacement of low- or high-density organic solutes. This study brings new insights into the energy cost and potential selective advantages of several strategies modulating the buoyancy of aflagellate marine phytoplankton.

A comparative study on edible Agaricus mushrooms as functional foods.

Agaricus bisporus is a cultivated mushroom; A. bitorquis, A. campestris and A. macrosporus are edible mushrooms growing wild in nature. A chemical characterization was carried out with samples that originated in Serbia. Antioxidant, antimicrobial and anti-quorum sensing properties of their methanolic and ethanolic extracts were assessed. A. campestris had the lowest caloric value and total sugar content and showed the highest concentration in organic and phenolic acids, as also in tocopherols (mainly γ-tocopherol). In general, the methanolic extracts showed higher antioxidant, but lower antibacterial and antifungal potential than ethanolic ones. Sub-inhibitory concentrations of the ethanolic extracts demonstrated reduction of virulence factors, AQ inhibition zones, twitching and swimming motility. The biofilm forming capability of P. aeruginosa PAO1 was also reduced in a concentration-dependent manner at sub-MIC values. The extracts of the tested Agaricus species are a promising source of antioxidant, antimicrobial and antiquorum sensing compounds.

Assessment of the optimal vitrification protocol for pre-pubertal mice testes leading to successful in vitro production of flagellated spermatozoa.

Testicular tissue cryopreservation offers the hope of preserved future fertility to pre-pubertal boys with cancer before exposition to gonadotoxic treatments. The objective of this study was to compare controlled slow freezing (CSF) with five vitrification techniques for cryopreservation of murine pre-pubertal testicular tissue and to evaluate the best protocol that could provide a successful completion of spermatogenesis after in vitro maturation. Testicular tissue from 24 mice at 6.5 days post-partum (dpp) was used to compare several vitrification protocols with one another, as well as with a CSF protocol. Toxicity test using additional 12 mice was performed for all cryopreservation solutions. Fresh tissue (FT) from six mice was used as a control. Once the optimal vitrification protocol was selected [the modified solid surface vitrification No. 1 (mSSV1 )], testes from 18 mice were cultured in vitro for 30 days with (i) fresh, (ii) slow-frozen/thawed and (iii) vitrified/warmed tissues. Testes from six mice at 36.5 dpp were used as controls. At day 30 of in vitro culture, germ cells of the seminiferous tubules showed a high ability to proliferate and elongated spermatids were observed after both freezing techniques, confirming the successful completion of in vitro spermatogenesis. However, after mSSV1 , the morphological alterations and the percentage of pyknotic seminiferous tubules were lower than CSF (4.67 ± 0.53 vs. 10.1 ± 1.12 and 22.7 ± 2.83% vs. 37.3 ± 4.24% respectively). Moreover, the number of flagellated spermatozoa produced per mg of tissue was higher for mSSV1 than for CSF (35 ± 3 vs. 9 ± 4 cells), with amounts of secreted testosterone during the culture close to those of FT. The mSSV1 protocol resulted in success rates better than CSF in maintaining testicular tissue structure, tubular morphology and tissue functions not solely for immediate frozen/thawed tissues but also after a long-term in vitro culture.

Flagellar filament bio-templated inorganic oxide materials - towards an efficient lithium battery anode.

Designing a new generation of energy-intensive and sustainable electrode materials for batteries to power a variety of applications is an imperative task. The use of biomaterials as a nanosized structural template for these materials has the potential to produce hitherto unachievable structures. In this report, we have used genetically modified flagellar filaments of the extremely halophilic archaea species Halobacterium salinarum to synthesize nanostructured iron oxide composites for use as a lithium-ion battery anode. The electrode demonstrated a superior electrochemical performance compared to existing literature results, with good capacity retention of 1032 mAh g(-1) after 50 cycles and with high rate capability, delivering 770 mAh g(-1) at 5 A g(-1) (~5 C) discharge rate. This unique flagellar filament based template has the potential to provide access to other highly structured advanced energy materials in the future.

The metabolic cost of flagellar motion in Pseudomonas putida KT2440.

Although the flagellar machinery of environmental bacteria endows cells with a phenomenal survival device, it also consumes much of the metabolic currency necessary for fuelling such a vigorous nano-motor. The physiological cost of flagella-related functions of the soil bacterium Pseudomonas putida KT2440 was examined and quantified through the deletion of a ≈ 70 kb DNA segment of the genome (≈ 1.1%), which includes relevant structural and regulatory genes in this micro-organism. The resulting strain lacked the protruding polar cords that define flagella in the wild-type P. putida strain and was unable of any swimming motility while showing a significant change in surface hydrophobicity. However, these deficiencies were otherwise concomitant with clear physiological advantages: rapid adaptation of the deleted strain to both glycolytic and gluconeogenic carbon sources, increased energy charge and, most remarkably, improved tolerance to oxidative stress, reflecting an increased NADPH/NADP(+) ratio. These qualities improve the endurance of non-flagellated cells to the metabolic fatigue associated with rapid growth in rich medium. Thus, flagellar motility represents the archetypal tradeoff involved in acquiring environmental advantages at the cost of a considerable metabolic burden.

Modeling the bacterial flagellum by an elastic network of rigid bodies.

Bacteria such as Escherichia coli propel themselves by rotating a bundle of helical filaments, each driven by a rotary motor embedded in the cell membrane. Each filament is an assembly of thousands of copies of the protein flagellin which assumes two different states. We model the filament by an elastic network of rigid bodies that form bonds with one another according to a scheme suggested by Namba and Vondervistz (1997 Q. Rev. Biophys. 30 1-65) and add additional binding sites at the inner part of the rigid body. Our model reproduces the helical parameters of the 12 possible polymorphic configurations very well. We demonstrate that its energetical ground state corresponds to the normal helical form, usually observed in nature, only when inner and outer binding sites of the rigid body have a large axial displacement. This finding correlates directly to the elongated shape of the flagellin molecule. An Ising Hamiltonian in our model directly addresses the two states of the flagellin protein. It contains an external field that represents external parameters which allow us to alter the ground state of the filament.

High-resolution x-ray observation of unstained samples by a newly developed SGXM.

Analytical methods with nanometre-scale resolution are indispensable in various scientific fields, including biology, chemistry and nanotechnology. One suitable tool, the soft x-ray microscope, provides high spatial resolution of unstained specimens. Recently, we developed an x-ray microscope called a scanning electron generation x-ray microscope (SGXM). This system had attached additional x-ray detection equipment to the scanning electron microscope (SEM). Here, we describe a new sample stage containing an improved x-ray emission film that enables the acquisition of high-resolution x-ray images of unstained samples. The method proposed in this study, which uses the new SGXM, enables us to acquire x-ray images by using an intact, conventional SEM without additional x-ray detection equipment. Moreover, experiments demonstrate that the new SGXM achieves a spatial resolution of 11 nm. Therefore, our system can be easily used for acquiring high-resolution images of a variety of samples across a broad range of scientific fields.

SALMATcor: microagglutination for Salmonella flagella serotyping.

Salmonella is a complex bacterial group with more than 2400 serovars widely distributed in nature; they are considered zoonotic because they can infect a variety of animals and be transmitted to humans. Usually, they cause alimentary acquired diseases such as gastroenteritis, typhoid fever, and others that can lead to severe complications and death. Serotyping is useful to differentiate among Salmonella, because it shows an important correlation with their clinical and epidemiological patterns; consequently, it is of high value for public health, animal health, agriculture, and industry. To characterize all known Kauffmann-White Salmonella serovars, over 250 antisera are required. Due to this and to high prices antisera, many laboratories worldwide have limitations in establishing Salmonella surveillance. Therefore, we developed and validated a Salmonella flagella microagglutination test (SALMATcor) that significantly reduces laboratory requirements of antisera. SALMATcor is based on scaling down, by fivefold, the antigen:antiserum volumes actually required for the reference method: flagella standard tube agglutination technique (STAT). Antigen preparation, temperatures, and incubation periods remained as established for STAT. The SALMATcor was validated according to ISO/DIS 16140:1999 protocol, which included 1187 comparisons of flagella determinations conducted by SALMATcor and STAT, on 141 Salmonella isolates of 12 common serotypes and the use of antiserum recommended for STAT. SALMATcor concordance was excellent (Cohen's kappa index 0.9982), obtaining relative accuracy >99.9% and relative specificity >99.9%. Additionally, SALMATcor has been used by CNRB-INCIENSA since 2004 to respond to all 40 Salmonella proficiency testing strains, provided by World Health Organization-Global Salmonella Surveillance Network, obtaining 100% concordance on serovar identification. On the basis of the results achieved with SALMATcor and considering that it also significantly reduces antiserum expenses, hand labor, glassware, and bench top and water bath space requirements (microtiter plates and micropipette tips are the only additional supplies), we envision that SALMATcor will contribute to establish a sustainable Salmonella serovar surveillance worldwide.