Antigen-specific acquired immunity in human brucellosis: implications for diagnosis, prognosis, and vaccine development.

Brucella spp., are Gram negative bacteria that cause disease by growing within monocyte/macrophage lineage cells. Clinical manifestations of brucellosis are immune mediated, not due to bacterial virulence factors. Acquired immunity to brucellosis has been studied through observations of naturally infected hosts (cattle, goats), mouse models (mice), and human infection. Even though Brucella spp. are known for producing mechanisms that evade the immune system, cell-mediated immune responses drive the clinical manifestations of human disease after exposure to Brucella species, as high antibody responses are not associated with protective immunity. The precise mechanisms by which cell-mediated immune responses confer protection or lead to disease manifestations remain undefined. Descriptive studies of immune responses in human brucellosis show that TH(1) (interferon-γ-producing T cells) are associated with dominant immune responses, findings consistent with animal studies. Whether these T cell responses are protective, or determine the different clinical responses associated with brucellosis is unknown, especially with regard to undulant fever manifestations, relapsing disease, or are associated with responses to distinct sets of Brucella spp. antigens are unknown. Few data regarding T cell responses in terms of specific recognition of Brucella spp. protein antigens and peptidic epitopes, either by CD4+ or CD8+ T cells, have been identified in human brucellosis patients. Additionally because current attenuated Brucella vaccines used in animals cause human disease, there is a true need for a recombinant protein subunit vaccine for human brucellosis, as well as for improved diagnostics in terms of prognosis and identification of unusual forms of brucellosis. This review will focus on current understandings of antigen-specific immune responses induced Brucella peptidic epitopes that has promise for yielding new insights into vaccine and diagnostics development, and for understanding pathogenetic mechanisms of human brucellosis

In silico prediction of molecular mechanisms of toxicity mediated by the leptospiral PF07598 gene family-encoded virulence-modifying proteins

Mechanisms of leptospirosis pathogenesis remain unclear despite the identification of a number of potential leptospiral virulence factors. We recently demonstrated potential mechanisms by which the virulence-modifying (VM) proteins—defined as containing a Domain of Unknown function (DUF1561), encoded by the PF07598 gene family—found only in group 1 pathogenic Leptospira—might mediate the clinical pathogenesis of leptospirosis. VM proteins belongs to classical AB toxin paradigm though have a unique AB domain architecture, unlike other AB toxins such as diphtheria toxin, pertussis toxin, shiga toxin, or ricin toxin which are typically encoded by two or more genes and self-assembled into a multi-domain holotoxin. Leptospiral VM proteins are secreted R-type lectin domain-containing exotoxins with discrete N-terminal ricin B-like domains involved in host cell surface binding, and a C-terminal DNase/toxin domain. Here we use the artificial intelligence-based AlphaFold algorithm and other computational tools to predict and elaborate on details of the VM protein structure-function relationship. Comparative AlphaFold and CD-spectroscopy defined the consistent secondary structure (Helix and ß-sheet) content, and the stability of the functional domains were further supported by molecular dynamics simulation. VM proteins comprises distinctive lectic family (QxW)3 motifs, the Mycoplasma CARDS toxin (D3 domain, aromatic patches), C-terminal similarity with mammalian DNase I. In-silico study proposed that Gln412, Gln523, His533, Thr59 are the high binding energy or ligand binding residues plausibly anticipates in the functional activities. Divalent cation (Mg+2-Gln412) and phosphate ion (PO4]−3-Arg615) interaction further supports the functional activities driven by C-terminal domain. Computation-driven structure-function studies of VM proteins will guide experimentation towards mechanistic understandings of leptospirosis pathogenesis, which underlie development of new therapeutic and preventive measures for this devastating disease

Expression of sialic acids and other nonulosonic acids in Leptospira

AbstractBackgroundSialic acids are negatively charged nine carbon backbone sugars expressed on mammalian cell surfaces. Sialic acids are part of a larger family of nonulosonic acid (NulO) molecules that includes pseudaminic and legionaminic acids. Microbial expression of sialic acids and other nonulosonic acids has been shown to contribute to host-microbe interactions in a variety of contexts, including participation in colonization, immune subversion, and behaviors such as biofilm formation, autoagglutination and motility. Previous research has suggested that some spirochetes may also express these molecules.ResultsHere we use a combination of molecular tools to investigate the presence of NulO biosynthetic gene clusters among clinical and saprophytic isolates of the genus Leptospira. Polymerase chain reaction and Southern blotting suggested that a variety of leptospires encoded NulO biosynthetic pathways. High performance liquid chromatography and mass spectrometry analyses provided biochemical evidence that di-N-acetylated NulO molecules are expressed at relatively high levels by L. interrogans serovar Lai strain 55601, and at lower levels by L. alexanderi serovar Manhao and L. fainei serovar Hurstbridge. Endogenous expression of N-acetylneuraminic acid (Neu5Ac, the most common sialic acid) was documented in L. interrogans serovar Copenhageni strain L1-130. Neu5Ac biosynthesis is also supported by a unique gene fusion event resulting in an enzyme with an N-terminal N-acetylneuraminic acid synthase domain and a C-terminal phosphatase domain. This gene fusion suggests that L. interrogans uses a Neu5Ac biosynthetic pathway more similar to animals than to other bacteria. Analysis of the composition and phylogeny of putative NulO biosynthetic gene clusters in L. interrogans serovar Lai and serovar Copenhageni revealed that both strains have complete biosynthetic pathways for legionamimic acid synthesis, a molecule with the same stereochemistry as sialic acid. Lectin-based affinity purification of NulO-modified molecules, followed by mass spectrometric identification suggests post-translational modification of surface lipoproteins, including Loa22.ConclusionsLeptospira species encode NulO biosynthetic pathways and synthesize multiple NulO molecules including sialic acid. Additional studies are needed to clarify the exact context and functional significance of NulO expression. These findings have implications for immune evasion during systemic leptospirosis

Ex vivo innate immune cytokine signature of enhanced risk of relapsing brucellosis.

BackgroundBrucellosis, a zoonotic infection caused by one of the Gram-negative intracellular bacteria of the Brucella genus, is an ongoing public health problem in Perú. While most patients who receive standard antibiotic treatment recover, 5-40% suffer a brucellosis relapse. In this study, we examined the ex vivo immune cytokine profiles of recovered patients with a history of acute and relapsing brucellosis.Methodology/principal findingsBlood was taken from healthy control donors, patients with a history of acute brucellosis, or patients with a history of relapsing brucellosis. Peripheral blood mononuclear cells were isolated and remained in culture without stimulation or were stimulated with a panel of toll-like receptor agonists or heat-killed Brucella melitensis (HKBM) isolates. Innate immune cytokine gene expression and protein secretion were measured by quantitative real-time polymerase chain reaction and a multiplex bead-based immunoassay, respectively. Acute and relapse patients demonstrated consistently elevated cytokine gene expression and secretion levels compared to controls. Notably, these include: basal and stimulus-induced expression of GM-CSF, TNF-α, and IFN-γ in response to LPS and HKBM; basal secretion of IL-6, IL-8, and TNF-α; and HKBM or Rev1-induced secretion of IL-1β, IL-2, GM-CSF, IFN-Υ, and TNF-α. Although acute and relapse patients were largely indistinguishable by their cytokine gene expression profiles, we identified a robust cytokine secretion signature that accurately discriminates acute from relapse patients. This signature consists of basal IL-6 secretion, IL-1β, IL-2, and TNF-α secretion in response to LPS and HKBM, and IFN-γ secretion in response to HKBM.Conclusions/significanceThis work demonstrates that informative cytokine variations in brucellosis patients can be detected using an ex vivo assay system and used to identify patients with differing infection histories. Targeted diagnosis of this signature may allow for better follow-up care of brucellosis patients through improved identification of patients at risk for relapse

Asymptomatic Renal Colonization of Humans in the Peruvian Amazon by Leptospira

Leptospirosis is a bacterial disease commonly transmitted from animals to humans. The more than 200 types of spiral-shaped bacteria (spirochetes) in the genus Leptospira are classified as pathogenic, intermediately pathogenic, or saprophytic (meaning not causing infection in any mammal) based on their ability to cause disease and on genetic information. Unique among the spirochetes that infect humans, Leptospira live both in the environment (in surface waters and moist soils), and in mammals, where they cause chronic infection by colonizing kidney tubules. Infected animals are the source of human infection, but humans have not been systematically studied as chronic Leptospira carriers. In our study, we found that more than 5% of people (in fact, only women) in a rural Amazonian village, without clinical evidence of infection by Leptospira, were chronically colonized by the bacteria. Chronic infection was not associated with a detectable immune response against the spirochete. Pathogenic and intermediately pathogenic Leptospira caused asymptomatic, chronic kidney infections. Future work is needed to determine whether such chronic infection can lead to human-to-human transmission of leptospirosis, and whether subtle measures of kidney disease are associated with asymptomatic, long-term leptospiral infection

Infection of laboratory-colonized Anopheles darlingi mosquitoes by Plasmodium vivax.

Anopheles darlingi Root is the most important malaria vector in the Amazonia region of South America. However, continuous propagation of An. darlingi in the laboratory has been elusive, limiting entomological, genetic/genomic, and vector-pathogen interaction studies of this mosquito species. Here, we report the establishment of an An. darlingi colony derived from wild-caught mosquitoes obtained in the northeastern Peruvian Amazon region of Iquitos in the Loreto Department. We show that the numbers of eggs, larvae, pupae, and adults continue to rise at least to the F6 generation. Comparison of feeding Plasmodium vivax ex vivo of F4 and F5 to F1 generation mosquitoes showed the comparable presence of oocysts and sporozoites, with numbers that corresponded to blood-stage asexual parasitemia and gametocytemia, confirming P. vivax vectorial capacity in the colonized mosquitoes. These results provide new avenues for research on An. darlingi biology and study of An. darlingi-Plasmodium interactions

Risk factors for diagnosed noma in northwest Nigeria: A case-control study, 2017

Background Noma (cancrum oris), a neglected tropical disease, rapidly disintegrates the hard and soft tissue of the face and leads to severe disfiguration and high mortality. The disease is poorly understood. We aimed to estimate risk factors for diagnosed noma to better guide existing prevention and treatment strategies using a case-control study design. Methods Cases were patients admitted between May 2015 and June 2016, who were under 15 years of age at reported onset of the disease. Controls were individuals matched to cases by village, age and sex. Caretakers answered the questionnaires. Risk factors for diagnosed noma were estimated by calculating unadjusted and adjusted odds ratios (ORs) and respective 95% confidence intervals (CI) using conditional logistic regression. Findings We included 74 cases and 222 controls (both median age 5 (IQR 3, 15)). Five cases (6.5%) and 36 (16.2%) controls had a vaccination card (p = 0.03). Vaccination coverage for polio and measles was below 7% in both groups. The two main reported water sources were a bore hole in the village (cases n = 27, 35.1%; controls n = 63, 28.4%; p = 0.08), and a well in the compound (cases n = 24, 31.2%; controls n = 102, 45.9%; p = 0.08). The adjusted analysis identified potential risk and protective factors for diagnosed noma which need further exploration. These include the potential risk factor of the child being fed pap every day (OR 9.8; CI 1.5, 62.7); and potential protective factors including the mother being the primary caretaker (OR 0.08; CI 0.01, 0.5); the caretaker being married (OR 0.006; CI 0.0006, 0.5) and colostrum being given to the baby (OR 0.4; CI 0.09, 2.09). Interpretation This study suggests that social conditions and infant feeding practices are potentially associated with being a diagnosed noma case in northwest Nigeria; these findings warrant further investigation into these factors