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Over recent decades, anthropogenic forest fires have significantly altered vegetation dynamics in the Amazon region. While human activities primarily initiate these fires, their escalation is intricately linked to climatic conditions, particularly droughts induced by the warm El Niño phase. This study investigates the impact of meteorological and hydrological drought on forest fires in the Amazon, focusing on the role of groundwater and El Niño events. Utilizing comprehensive drought indicators at various soil depths and standardized precipitation indexes, the research spans from 2004 to 2016, revealing a consistent decrease in humidity conditions across surface soil moisture, root zone soil moisture, and groundwater storage levels. With its slower response to precipitation changes, groundwater emerges as a crucial factor influencing hydrological drought patterns in the Amazon. The spatial distribution of drought conditions is explored, highlighting areas with lower humidity concentrations in the northeast and a correlation between forest fires and positive rates of change in burned area fraction during El Niño events. Notably, the study underscores the substantial increase in burned area during the 2015-2016, characterized by a very strong El Niño. This nuanced understanding of groundwater dynamics and its interplay with El Niño events provides critical insights for developing a tailored fire risk index in the ecologically significant and vulnerable Amazon basin, subsidizing strategies for mitigating fire risk and enhancing preparedness.

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Drought is considered the most severe water-related disaster in the Cauto river basin, which is the longest river and the main agricultural producer in Cuba. Better understanding of drought characteristics is crucial to drought management. Given the sparsity of ground-based precipitation observations in the Cauto, this study aims at using gridded global precipitation to analyze the spatio-temporal variations of drought in this river basin. Firstly, the monthly Climate Hazards Group InfraRed Precipitation with Station data (CHIRPS) was calibrated with the gauged precipitation using the Thiessen polygon-based method and linear least squares regression equations. Then, the gridded standardized precipitation index (SPI) with time scales of 3, 6, 9 months and drought characteristics, namely, drought frequency, duration and intensity were calculated using the calibrated CHIRPS. Finally, the spatio-temporal analysis was performed to investigate the variations of drought in the Cauto river basin in time and space. The obtained results show that the calibrated CHIRPS is highly consistent with the gauged observations and is capable of determining the magnitude, time, and spatial extent of drought events in the Cauto river basin. The trend analysis by the Mann-Kendall test reveals that although the trend is not statistically significant, the SPI tends to decrease with time in the dry season, which indicates the more severe drought. The spatial analysis indicates that the lower altitude area of the Cauto river basin is suffered from longer drought duration and higher drought intensity than the upper one. This study expresses the importance of open global precipitation data sources in monitoring and quantifying drought characteristics in data-scarce regions.

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Excisable genomic islands (EGIs) are horizontally acquired genetic elements that harbor an array of genes with diverse functions. ROD21 is an EGI found integrated in the chromosome of Salmonella enterica serovar Enteritidis (Salmonella ser. Enteritidis). While this island is known to be involved in the capacity of Salmonella ser. Enteritidis to cross the epithelial barrier and colonize sterile organs, the role of most ROD21 genes remains unknown, and thus, the identification of their function is fundamental to understanding the impact of this EGI on bacterium pathogenicity. Therefore, in this study, we used a bioinformatical approach to evaluate the function of ROD21-encoded genes and delve into the characterization of SEN1990, a gene encoding a putative DNA-binding protein. We characterized the predicted structure of SEN1990, finding that this protein contains a three-stranded winged helix-turn-helix (wHTH) DNA-binding domain. Additionally, we identified homologs of SEN1990 among other members of the EARL EGIs. Furthermore, we deleted SEN1990 in Salmonella ser. Enteritidis, finding no differences in the replication or maintenance of the excised ROD21, contrary to what the previous Refseq annotation of the protein suggests. High-throughput RNA sequencing was carried out to evaluate the effect of the absence of SEN1990 on the bacterium's global transcription. We found a downregulated expression of oafB, an SPI-17-encoded acetyltransferase involved in O-antigen modification, which was restored when the deletion mutant was complemented ectopically. Additionally, we found that strains lacking SEN1990 had a reduced capacity to colonize sterile organs in mice. Our findings suggest that SEN1990 encodes a wHTH domain-containing protein that modulates the transcription of oafB from the SPI-17, implying a crosstalk between these pathogenicity islands and a possible new role of ROD21 in the pathogenesis of Salmonella ser. Enteritidis.

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The type VI secretion system (T6SS) is a contact-dependent contractile multiprotein apparatus widely distributed in Gram-negative bacteria. These systems can deliver different effector proteins into target bacterial and/or eukaryotic cells, contributing to the environmental fitness and virulence of many bacterial pathogens. Salmonella harbors five different T6SSs encoded in different genomic islands. The T6SS encoded in Salmonella Pathogenicity Island 6 (SPI-6) contributes to Salmonella competition with the host microbiota and its interaction with infected host cells. Despite its relevance, information regarding the total number of effector proteins encoded within SPI-6 and its distribution among different Salmonella enterica serotypes is limited. In this work, we performed bioinformatic and comparative genomics analyses of the SPI-6 T6SS gene cluster to expand our knowledge regarding the T6SS effector repertoire and the global distribution of these effectors in Salmonella. The analysis of a curated dataset of 60 Salmonella enterica genomes from the Secret6 database revealed the presence of 23 new putative T6SS effector/immunity protein (E/I) modules. These effectors were concentrated in the variable regions 1 to 3 (VR1-3) of the SPI-6 T6SS gene cluster. VR1-2 were enriched in candidate effectors with predicted peptidoglycan hydrolase activity, while VR3 was enriched in candidate effectors of the Rhs family with C-terminal extensions with predicted DNase, RNase, deaminase, or ADP-ribosyltransferase activity. A global analysis of known and candidate effector proteins in Salmonella enterica genomes from the NCBI database revealed that T6SS effector proteins are differentially distributed among Salmonella serotypes. While some effectors are present in over 200 serotypes, others are found in less than a dozen. A hierarchical clustering analysis identified Salmonella serotypes with distinct profiles of T6SS effectors and candidate effectors, highlighting the diversity of T6SS effector repertoires in Salmonella enterica. The existence of different repertoires of effector proteins suggests that different effector protein combinations may have a differential impact on the environmental fitness and pathogenic potential of these strains.

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In many bacteria, the BarA/SirA and Csr regulatory systems control expression of genes encoding a wide variety of cellular functions. The BarA/SirA two-component system induces the expression of CsrB and CsrC, two small non-coding RNAs that sequester CsrA, a protein that binds to target mRNAs and thus negatively or positively regulates their expression. BarA/SirA and CsrB/C induce expression of the Salmonella Pathogenicity Island 1 (SPI-1) genes required for Salmonella invasion of host cells. To further investigate the regulatory role of the BarA/SirA and Csr systems in Salmonella, we performed LC-MS/MS proteomic analysis using the WT S. Typhimurium strain and its derived ΔsirA and ΔcsrB ΔcsrC mutants grown in SPI-1-inducing conditions. The expression of 164 proteins with a wide diversity, or unknown, functions was significantly affected positively or negatively by the absence of SirA and/or CsrB/C. Interestingly, 19 proteins were identified as new targets for SirA-CsrB/C. Our results support that SirA and CsrB/C act in a cascade fashion to regulate gene expression in S. Typhimurium in the conditions tested. Notably, our results show that SirA-CsrB/C-CsrA controls expression of proteins required for the replication of Salmonella in the intestinal lumen, in an opposite way to its control exerted on the SPI-1 proteins. SIGNIFICANCE: The BarA/SirA and Csr global regulatory systems control a wide range of cellular processes, including the expression of virulence genes. For instance, in Salmonella, BarA/SirA and CsrB/C positively regulate expression of the SPI-1 genes, which are required for Salmonella invasion to host cells. In this study, by performing a proteomic analysis, we identified 164 proteins whose expression was positively or negatively controlled by SirA and CsrB/C in SPI-1-inducing conditions, including 19 new possible targets of these systems. Our results support the action of SirA and CsrB/C in a cascade fashion to control different cellular processes in Salmonella. Interestingly, our data indicate that SirA-CsrB/C-CsrA controls inversely the expression of proteins required for invasion of the intestinal epithelium and for replication in the intestinal lumen, which suggests a role for this regulatory cascade as a molecular switch for Salmonella virulence. Thus, our study further expands the insight into the regulatory mechanisms governing the virulence and physiology of an important pathogen.

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Salmonella genus harbors five Type VI Secretion System (T6SS) gene clusters. The T6SS encoded in SPI-6 (T6SSSPI-6) contributes to Salmonella Typhimurium colonization of chickens and mice, while the T6SS encoded in SPI-19 (T6SSSPI-19) of Salmonella Gallinarum contributes to chicken colonization. Interestingly, the T6SSSPI-19 of Salmonella Gallinarum complemented the defect in chicken colonization of a Salmonella Typhimurium strain that lacks the T6SSSPI-6, suggesting that both T6SSs are interchangeable. Here we show that the transfer of Salmonella Gallinarum T6SSSPI-19 complemented the defect in mice colonization of a Salmonella Typhimurium ΔT6SSSPI-6 strain, indicating that both T6SSs are functionally redundant during host colonization.

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An integrative approach combining genomics, transcriptomics, and cell biology is presented to address leaf scald disease, a major problem for the sugarcane industry. To gain insight into the biology of the causal agent, the complete genome sequences of four Brazilian Xanthomonas albilineans strains with differing virulence capabilities are presented and compared to the GPEPC73 reference strain and FJ1. Based on the aggressiveness index, different strains were compared: Xa04 and Xa11 are highly aggressive, Xa26 is intermediate, and Xa21 is the least, while, based on genome structure, Xa04 shares most of its genomic features with Xa26, and Xa11 share most of its genomic features with Xa21. In addition to presenting more clustered regularly interspaced short palindromic repeats (CRISPR) clusters, four more novel prophage insertions are present than the previously sequenced GPEPC73 and FJ1 strains. Incorporating the aggressiveness index and in vitro cell biology into these genome features indicates that disease establishment is not a result of a single determinant factor, as in most other Xanthomonas species. The Brazilian strains lack the previously described plasmids but present more prophage regions. In pairs, the most virulent and the least virulent share unique prophages. In vitro transcriptomics shed light on the 54 most highly expressed genes among the 4 strains compared to ribosomal proteins (RPs), of these, 3 outer membrane proteins. Finally, comparative albicidin inhibition rings and in vitro growth curves of the four strains also do not correlate with pathogenicity. In conclusion, the results disclose that leaf scald disease is not associated with a single shared characteristic between the most or the least pathogenic strains. IMPORTANCE An integrative approach is presented which combines genomics, transcriptomics, and cell biology to address leaf scald disease. The results presented here disclose that the disease is not associated with a single shared characteristic between the most pathogenic strains or a unique genomic pattern. Sequence data from four Brazilian strains are presented that differ in pathogenicity index: Xa04 and Xa11 are highly virulent, Xa26 is intermediate, and Xa21 is the least pathogenic strain, while, based on genome structure, Xa04 shares with Xa26, and Xa11 shares with X21 most of the genome features. Other than presenting more CRISPR clusters and prophages than the previously sequenced strains, the integration of aggressiveness and cell biology points out that disease establishment is not a result of a single determinant factor as in other xanthomonads.

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Drought indices are a numerical representation of drought conditions aimed to provide quantitative assessments of the magnitude, spatial extent, timing, and duration of drought events. Since the adverse effects of droughts vary according to the characteristics of the event, the socioeconomic vulnerabilities, exposed communities or environments, there is a profusion of drought indicators to assess drought impacts in different sectors. In this study, we evaluated the performance of two drought indices, the Standardized Precipitation Index-SPI and Standardized Precipitation Evapotranspiration Index-SPEI over Brazil derived from gridded meteorological information over the period 1980-2019. Firstly, we compared the gridded derived indices against the same indices derived from weather station data and available from a global dataset for time scales of 3, 6, 12, 24 months. Then we analyzed the spatio-temporal trends in SPI and SPEI time-series, which revealed statistically significant trends toward drier conditions across central Brazil for all time scales, though with more intensity for time scales of 12 months and larger. Trends were more significant in magnitude for SPEI than SPI, indicating an important role in the increase in evaporation, driven by increasingly higher temperatures. Finally, we demonstrated that climate signals are already having a disruptive effect on the country's energy security. Supplementary Information: The online version contains supplementary material available at 10.1007/s11069-022-05759-0.

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Cinnamaldehyde (CNM) is a cyclic terpene alcohol found as the major compound of essential oils from some plants of the genus Cinnamomum (Lauraceae). CNM has several reported pharmacological activities, including antimicrobial, antivirulence, antioxidant, and immunomodulatory effects. These properties make CNM an attractive lead molecule for the development of anti-infective agents. In this descriptive review, we discuss the application of CNM in experimental models of microbial infection using invertebrate and vertebrate organisms. CNM (pure or in formulations) has been successfully applied in the treatment of infections caused by a range of bacterial (such as Cronobacter sakazakii, Escherichia coli, Listeria monocytogenes, Mycobacterium tuberculosis, Pseudomonas aeruginosa, Salmonella enterica, Staphylococcus aureus, Streptococcus agalactiae, Vibrio cholerae) and fungal (such as Aspergillus fumigatus, Candida albicans and Cryptococcus neoformans) pathogens. All these experimental evidence-based findings have promoted the use of cinnamaldehyde as the leading molecule for developing new anti- infective drugs.

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The acquisition of Salmonella pathogenicity island 2 (SPI-2) conferred on Salmonella the ability to survive and replicate within host cells. The ssrAB bicistronic operon, located in SPI-2, encodes the SsrAB two-component system (TCS), which is the central positive regulator that induces the expression of SPI-2 genes as well as other genes located outside this island. On the other hand, CpxRA is a two-component system that regulates expression of virulence genes in many bacteria in response to different stimuli that perturb the cell envelope. We previously reported that the CpxRA system represses the expression of SPI-1 and SPI-2 genes under SPI-1-inducing conditions by decreasing the stability of the SPI-1 regulator HilD. Here, we show that under SPI-2-inducing conditions, which mimic the intracellular environment, CpxRA represses the expression of SPI-2 genes by the direct action of phosphorylated CpxR (CpxR-P) on the ssrAB regulatory operon. CpxR-P recognized two sites located proximal and distal from the promoter located upstream of ssrA. Consistently, we found that CpxRA reduces the replication of Salmonella enterica serovar Typhimurium inside murine macrophages. Therefore, our results reveal CpxRA as an additional regulator involved in the intracellular lifestyle of Salmonella, which in turn adds a new layer to the intricate regulatory network controlling the expression of Salmonella virulence genes. IMPORTANCE SPI-2 encodes a type III secretion system (T3SS) that is a hallmark for the species Salmonella enterica, which is essential for the survival and replication within macrophages. Expression of SPI-2 genes is positively controlled by the two-component system SsrAB. Here, we determined a regulatory mechanism involved in controlling the overgrowth of Salmonella inside macrophages. In this mechanism, CpxRA, a two-component system that is activated by extracytoplasmic stress, directly represses expression of the ssrAB regulatory operon; as a consequence, expression of SsrAB target genes is decreased. Our findings reveal a novel mechanism involved in the intracellular lifestyle of Salmonella, which is expected to sense perturbations in the bacterial envelope that Salmonella faces inside host cells, as the synthesis of the T3SS-2 itself.

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In the last decade, the vision systems have improved their capabilities to capture 3D images in bad weather scenarios. Currently, there exist several techniques for image acquisition in foggy or rainy scenarios that use infrared (IR) sensors. Due to the reduced light scattering at the IR spectra it is possible to discriminate the objects in a scene compared with the images obtained in the visible spectrum. Therefore, in this work, we proposed 3D image generation in foggy conditions using the single-pixel imaging (SPI) active illumination approach in combination with the Time-of-Flight technique (ToF) at 1550 nm wavelength. For the generation of 3D images, we make use of space-filling projection with compressed sensing (CS-SRCNN) and depth information based on ToF. To evaluate the performance, the vision system included a designed test chamber to simulate different fog and background illumination environments and calculate the parameters related to image quality.

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One important event for the divergence of Salmonella from Escherichia coli was the acquisition by horizontal transfer of the Salmonella pathogenicity island 1 (SPI-1), containing genes required for the invasion of host cells by Salmonella. HilD is an AraC-like transcriptional regulator in SPI-1 that induces the expression of the SPI-1 and many other acquired virulence genes located in other genomic regions of Salmonella. Additionally, HilD has been shown to positively control the expression of some ancestral genes (also present in E. coli and other bacteria), including phoH. In this study, we determined that both the gain of HilD and cis-regulatory evolution led to the integration of the phoH gene into the HilD regulon. Our results indicate that a HilD-binding sequence was generated in the regulatory region of the S. enterica serovar Typhimurium phoH gene, which mediates the activation of promoter 1 of this gene under SPI-1-inducing conditions. Furthermore, we found that repression by H-NS, a histone-like protein, was also adapted on the S. Typhimurium phoH gene and that HilD activates the expression of this gene in part by antagonizing H-NS. Additionally, our results revealed that the expression of the S. Typhmurium phoH gene is also activated in response to low phosphate but independently of the PhoB/R two-component system, known to regulate the E. coli phoH gene in response to low phosphate. Thus, our results indicate that cis-regulatory evolution has played a role in the expansion of the HilD regulon and illustrate the phenomenon of differential regulation of ortholog genes. IMPORTANCE Two mechanisms mediating differentiation of bacteria are well known: acquisition of genes by horizontal transfer events and mutations in coding DNA sequences. In this study, we found that the phoH ancestral gene is differentially regulated between Salmonella Typhimurium and Escherichia coli, two closely related bacterial species. Our results indicate that this differential regulation was generated by mutations in the regulatory sequence of the S. Typhimurium phoH gene and by the acquisition by S. Typhimurium of foreign DNA encoding the transcriptional regulator HilD. Thus, our results, together with those from an increasing number of studies, indicate that cis-regulatory evolution can lead to the rewiring and reprogramming of transcriptional regulation, which also plays an important role in the divergence of bacteria through time.

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Droughts threaten water resources, agriculture, socio-economic activities and the population at the global and regional level, so identifying areas with homogeneous drought behaviors is an important consideration in improving the management of water resources. The objective of this study is to identify homogenous zones over Paraíba State in relation to the state, duration and severity of droughts that have occurred over the last 20 years (1998-2017) using hierarchical cluster analysis based on both gauge-measured and Tropical Rainfall Measuring Mission (TRMM) estimated rainfall data (TMPA 3B42). The drought series were calculated using the Standardized Precipitation Index (SPI) based on eight time scales and were grouped according to drought state, duration and severity time series. The integrated results of state, duration and severity of droughts indicate that there is a basis for dividing Paraíba State into two major regions (a) Zone I, formed by Mata Paraibana and Agreste Paraibano, and (b) Zone II, composed by Borborema and Sertão Paraibano. This division is evident when assessing short-term droughts, but in the case of long-term droughts, Paraíba State has a high similarity in terms of drought state, duration, and severity. Factors such as proximity to the ocean, active climatic systems, and the local relief configuration were identified as influencing the drought regime. Finally, it is concluded that TMPA rainfall estimates represent a valuable source of data to regionalize and identify drought patterns over this part of Brazil and that other studies of this type should be carried out to monitor these phenomena based on other satellite-based rainfall data, including the Global Precipitation Mission (GPM).

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Salmonella enterica is a common source of food and water-borne infections, causing a wide range of clinical ailments in both human and animal hosts. Immunity to Salmonella involves an interplay between different immune responses, which are rapidly initiated to control bacterial burden. However, Salmonella has developed several strategies to evade and modulate the host immune responses. In this sense, the main knowledge about the pathogenicity of this bacterium has been obtained by the study of mouse models with non-typhoidal serovars. However, this knowledge is not representative of all the pathologies caused by non-typhoidal serovars in the human. Here we review the most important features of typhoidal and non-typhoidal serovars and the diseases they cause in the human host, describing the virulence mechanisms used by these pathogens that have been identified in different models of infection.

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Growing evidence indicates that small noncoding RNAs (sRNAs) play important regulatory roles during bacterial infection. In Salmonella Typhimurium, several sRNAs are strongly up-regulated within macrophages, but little is known about their role during the infection process. Among these sRNAs, the well-characterized paralogs RyhB-1 and RyhB-2 are two regulators of gene expression mainly related with the response to iron availability. To investigate the role of the sRNAs RyhB-1 and RyhB-2 from S. Typhimurium in the infection of RAW264.7 macrophages, we analyzed several phenotypic traits from intracellular mutant strains lacking one and both sRNAs. Deletion of RyhB-1 and/or RyhB-2 resulted in increased intracellular survival and faster replication within macrophages. The bacterial metabolic status inside macrophages was also analyzed, revealing that all the mutant strains exhibited higher intracellular levels of ATP and lower NAD+/NADH ratios than the wild type. Expression analyses from bacteria infecting macrophages showed that RyhB-1 and RyhB-2 affect the intra-macrophage expression of bacterial genes associated with the Salmonella pathogenicity island 1 (SPI-1) and the type III secretion system (T3SS). With a two-plasmid system and compensatory mutations, we confirmed that RyhB-1 and RyhB-2 directly interact with the mRNAs of the invasion chaperone SicA and the regulatory protein RtsB. Altogether, these results indicate that the RyhB homologs contribute to the S. Typhimurium virulence modulation inside macrophages by reducing the intracellular growth and down-regulating the SPI-1 gene expression.

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The type III secretion systems (T3SS) encoded in pathogenicity islands SPI-1 and SPI-2 are key virulence factors of Salmonella. These systems translocate proteins known as effectors into eukaryotic cells during infection. To characterize the functionality of T3SS effectors, gene fusions to the CyaA' reporter of Bordetella pertussis are often used. CyaA' is a calmodulin-dependent adenylate cyclase that is only active within eukaryotic cells. Thus, the translocation of an effector fused to CyaA' can be evaluated by measuring cAMP levels in infected cells. Here, we report the construction of plasmids pCyaA'-Kan and pCyaA'-Cam, which contain the ORF encoding CyaA' adjacent to a cassette that confers resistance to kanamycin or chloramphenicol, respectively, flanked by Flp recombinase target (FRT) sites. A PCR product from pCyaA'-Kan or pCyaA'-Cam containing these genetic elements can be introduced into the bacterial chromosome to generate gene fusions by homologous recombination using the Red recombination system from bacteriophage λ. Subsequently, the resistance cassette can be removed by recombination between the FRT sites using the Flp recombinase. As a proof of concept, the plasmids pCyaA'-Kan and pCyaA'-Cam were used to generate unmarked chromosomal fusions of 10 T3SS effectors to CyaA' in S. Typhimurium. Each fusion protein was detected by Western blot using an anti-CyaA' monoclonal antibody when the corresponding mutant strain was grown under conditions that induce the expression of the native gene. In addition, T3SS-1-dependent secretion of fusion protein SipA-CyaA' during in vitro growth was verified by Western blot analysis of culture supernatants. Finally, efficient translocation of SipA-CyaA' into HeLa cells was evidenced by increased intracellular cAMP levels at different times of infection. Therefore, the plasmids pCyaA'-Kan and pCyaA'-Cam can be used to generate unmarked chromosomal cyaA' translational fusion to study regulated expression, secretion and translocation of Salmonella T3SS effectors into eukaryotic cells.

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Type VI secretion systems (T6SSs) are nanomachines used by bacteria to inject toxic effectors into competitors. The identity and mechanism of many effectors remain unknown. We characterized a Salmonella T6SS antibacterial effector called Tlde1 that is toxic in target-cell periplasm and is neutralized by its cognate immunity protein (Tldi1). Microscopy analysis reveals that cells expressing Tlde1 stop dividing and lose cell envelope integrity. Bioinformatic analysis uncovers similarities between Tlde1 and the catalytic domain of l,d-transpeptidases. Point mutations on conserved catalytic residues abrogate toxicity. Biochemical assays reveal that Tlde1 displays both l,d-carboxypeptidase activity by cleaving peptidoglycan tetrapeptides between meso-diaminopimelic acid3 and d-alanine4 and l,d-transpeptidase exchange activity by replacing d-alanine4 by a non-canonical d-amino acid. Phylogenetic analysis shows that Tlde1 homologs constitute a family of T6SS-associated effectors broadly distributed among Proteobacteria. This work expands our current knowledge about bacterial effectors used in interbacterial competition and reveals a different mechanism of bacterial antagonism.

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BACKGROUND: Salmonella pathogenicity island (SPI)-13 is conserved in many serovars of S. enterica, including S. Enteritidis, S. Typhimurium and S. Gallinarum. However, it is absent in typhoid serovars such as S. Typhi and Paratyphi A, which carry SPI-8 at the same genomic location. Because the interaction with macrophages is a critical step in Salmonella pathogenicity, in this study we investigated the role played by SPI-13 and SPI-8 in the interaction of S. Enteritidis and S. Typhi with cultured murine (RAW264.7) and human (THP-1) macrophages. RESULTS: Our results showed that SPI-13 was required for internalization of S. Enteritidis in murine but not human macrophages. On the other hand, SPI-8 was not required for the interaction of S. Typhi with human or murine macrophages. Of note, the presence of an intact copy of SPI-13 in a S. Typhi mutant carrying a deletion of SPI-8 did not improve its ability to be internalized by, or survive in human or murine macrophages. CONCLUSIONS: Altogether, our results point out to different roles for SPI-13 and SPI-8 during Salmonella infection. While SPI-13 contributes to the interaction of S. Enteritidis with murine macrophages, SPI-8 is not required in the interaction of S. Typhi with murine or human macrophages. We hypothesized that typhoid serovars have lost SPI-13 and maintained SPI-8 to improve their fitness during another phase of human infection.

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BACKGROUND: Salmonella pathogenicity island (SPI)-13 is conserved in many serovars of S. enterica, including S. Enteritidis, S. Typhimurium and S. Gallinarum. However, it is absent in typhoid serovars such as S. Typhi and Paratyphi A, which carry SPI-8 at the same genomic location. Because the interaction with macrophages is a critical step in Salmonella pathogenicity, in this study we investigated the role played by SPI-13 and SPI-8 in the interaction of S. Enteritidis and S. Typhi with cultured murine (RAW264.7) and human (THP-1) macrophages. RESULTS: Our results showed that SPI-13 was required for internalization of S. Enteritidis in murine but not human macrophages. On the other hand, SPI-8 was not required for the interaction of S. Typhi with human or murine macrophages. Of note, the presence of an intact copy of SPI-13 in a S. Typhi mutant carrying a deletion of SPI-8 did not improve its ability to be internalized by, or survive in human or murine macrophages. CONCLUSIONS: Altogether, our results point out to different roles for SPI-13 and SPI-8 during Salmonella infection. While SPI-13 contributes to the interaction of S. Enteritidis with murine macrophages, SPI-8 is not required in the interaction of S. Typhi with murine or human macrophages. We hypothesized that typhoid serovars have lost SPI-13 and maintained SPI-8 to improve their fitness during another phase of human infection.

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The social amoeba Dictyostelium discoideum has proven to be a useful model for studying relevant aspects of the host-pathogen interaction. In this work, D. discoideum was used as a model to study the ability of Salmonella Typhimurium to survive in amoebae and to evaluate the contribution of selected genes in this process. To do this, we performed infection assays using axenic cultures of D. discoideum co-cultured with wild-type S. Typhimurium and/or defined mutant strains. Our results confirmed that wild-type S. Typhimurium is able to survive intracellularly in D. discoideum. In contrast, mutants ΔaroA and ΔwaaL are defective in intracellular survival in this amoeba. Next, we included in our study a group of mutants in genes directly linked to Salmonella virulence. Of note, mutants ΔinvA, ΔssaD, ΔclpV, and ΔphoPQ also showed an impaired ability to survive intracellularly in D. discoideum. This indicates that S. Typhimurium requires a functional biosynthetic pathway of aromatic compounds, a lipopolysaccharide containing a complete O-antigen, the type III secretion systems (T3SS) encoded in SPI-1 and SPI-2, the type VI secretion system (T6SS) encoded in SPI-6 and PhoP/PhoQ two-component system to survive in D. discoideum. To our knowledge, this is the first report on the requirement of O-antigen and T6SS in the survival of Salmonella within amoebae. In addition, mutants ΔinvA and ΔssaD were internalized in higher numbers than the wild-type strain during competitive infections, suggesting that S. Typhimurium requires the T3SS encoded in SPI-1 and SPI-2 to evade phagocytosis by D. discoideum. Altogether, these results indicate that S. Typhimurium exploits a common set of genes and molecular mechanisms to survive within amoeba and animal host cells. The use of D. discoideum as a model for host-pathogen interactions will allow us to discover the gene repertoire used by Salmonella to survive inside the amoeba and to study the cellular processes that are affected during infection.