Middle East Respiratory Syndrome Coronavirus Infection Elicits Long-lasting Specific Antibody, T and B Cell Immune Responses in Recovered Individuals.

BACKGROUND: The Middle East respiratory syndrome coronavirus (MERS-CoV) is a highly pathogenic zoonotic betacoronavirus and a global public health concern. Better undersetting of the immune responses to MERS-CoV is needed to characterize the correlates of protection and durability of the immunity and to aid in developing preventative and therapeutic interventions. Although MERS-CoV-specific circulating antibodies could persist for several years post-recovery, their waning raises concerns about their durability and role in protection. Nonetheless, memory B and T cells could provide long-lasting protective immunity despite the serum antibodies levels. METHODS: Serological and flow cytometric analysis of MERS-CoV-specific immune responses were performed on samples collected from a cohort of recovered individuals who required intensive care unit (ICU) admission as well as hospital or home isolation several years after infection to characterize the longevity and quality of humoral and cellular immune responses. RESULTS: Our data showed that MERS-CoV infection could elicit robust long-lasting virus-specific binding and neutralizing antibodies as well as T- and B-cell responses up to 6.9 years postinfection regardless of disease severity or need for ICU admission. Apart from the persistent high antibody titers, this response was characterized by B-cell subsets with antibody-independent functions as demonstrated by their ability to produce tumor necrosis factor α (TNF-α), interleukin (IL)-6, and interferon γ (IFN-γ) cytokines in response to antigen stimulation. Furthermore, virus-specific activation of memory CD8+ and CD4+ T cell subsets from MERS-recovered patients resulted in secretion of high levels of TNF-α, IL-17, and IFN-γ. CONCLUSIONS: MERS-CoV infection could elicit robust long-lasting virus-specific humoral and cellular responses.

Anti-α-Gal antibodies detected by novel neoglycoproteins as a diagnostic tool for Old World cutaneous leishmaniasis caused by Leishmania major.

Outbreaks of Old World cutaneous leishmaniasis (CL) have significantly increased due to the conflicts in the Middle East, with most of the cases occurring in resource-limited areas such as refugee settlements. The standard methods of diagnosis include microscopy and parasite culture, which have several limitations. To address the growing need for a CL diagnostic that can be field applicable, we have identified five candidate neoglycoproteins (NGPs): Galα (NGP3B), Galα(1,3)Galα (NGP17B), Galα(1,3)Galß (NGP9B), Galα(1,6)[Galα(1,2)]Galß (NGP11B), and Galα(1,3)Galß(1,4)Glcß (NGP1B) that are differentially recognized in sera from individuals with Leishmania major infection as compared with sera from heterologous controls. These candidates contain terminal, non-reducing α-galactopyranosyl (α-Gal) residues, which are known potent immunogens to humans. Logistic regression models found that NGP3B retained the best diagnostic potential (area under the curve from receiver-operating characteristic curve = 0.8). Our data add to the growing body of work demonstrating the exploitability of the human anti-α-Gal response in CL diagnosis.

Detection of high levels of anti-α-galactosyl antibodies in sera of patients with Old World cutaneous leishmaniasis: a possible tool for diagnosis and biomarker for cure in an elimination setting.

In the Kingdom of Saudi Arabia (KSA), Old World cutaneous leishmaniasis (CL) is mainly caused by Leishmania major and Leishmania tropica parasites. Diagnosis of CL is predominately made by clinicians, who at times fail to detect the disease and are unable to identify parasite species. Here, we report the development of a chemiluminescent enzyme-linked immunosorbent assay (CL-ELISA) to measure the levels of anti-α-galactosyl antibodies in human sera. Using this assay, we have found that individuals infected with either Leishmania spp. had significantly elevated levels (up to 9-fold higher) of anti-α-Gal IgG compared to healthy control individuals. The assay sensitivity was 96% for L. major (95% CI; 94-98%) and 91% for L. tropica (95% CI; 86-98%) infections and therefore equivalent to restriction fragment length polymorphism-polymerase chain reaction analysis of parasite ITS1 gene. In addition, the assay had higher sensitivity than microscopy analysis, which only detected 68 and 45% of the L. major and L. tropica infections, respectively. Interestingly, up to 2 years following confirmed CL cure individuals had 28-fold higher levels of anti-α-Gal IgG compared to healthy volunteers. Monitoring levels of anti-α-Gal antibodies can be exploited as both a diagnostic tool and as a biomarker of cure of Old World CL in disease elimination settings.

Chromosome-Scale Assembly of the Complete Genome Sequence of Leishmania (Mundinia) procaviensis Isolate 253, Strain LV425.

Leishmania (Mundinia) procaviensis is a parasitic kinetoplastid that was first isolated from a rock hyrax in Namibia in 1975. We present the complete genome sequence of Leishmania (Mundinia) procaviensis isolate 253, strain LV425, sequenced using combined short- and long-read technologies. This genome will contribute to our understanding of hyraxes as a Leishmania reservoir.

Isolation, identification and antimicrobial susceptibility of the bacteria isolated from Hyalomma dromedarii infesting camels in Al-Jouf province, Saudi Arabia.

Ticks are important ectoparasites that transmit various pathogens causing morbidity and mortality in humans and animals. Saudi Arabia faces several challenges that can contribute to the emergence and spread of antimicrobial resistance (AMR) bacteria. These challenges require collaborative efforts to successfully achieve significant control of AMR in the country. The present study aims to isolate bacteria from camels' tick Hyalomma dromedarii in Al-Jouf province to identify and determine these isolates' antimicrobial susceptibilities. Forty-nine ticks were collected from dromedary camels and morphologically classified as H. dromedarii. Ticks were then homogenized and plated individually, which resulted in the isolation of 55 bacteria. The results showed that the bacterial isolates belong to 20 different species. About 71% (n = 39) of the total isolates were identified as Gram-positive bacteria comprised of 11 different species, while 29% (n = 16) of the total isolates were Gram-negative bacteria comprised of 9 different species. The most prevalent isolate within the total samples was Staphylococcus lentus (22.45%, 11/49), followed by Staphylococcus pseudintermedius (18.37%, 9/49) and Sphingomonas paucimobilis (16.33% 8/49). The antimicrobial susceptibility profile of Gram-positive bacteria showed that 100% (n = 31) were resistant to benzylpenicillin; 90.3% (n = 28) were resistant to oxacillin; 58.1% (n = 18) were resistant to clindamycin; 48.4% (n = 15) were resistant to vancomycin. In addition, 32.3% (n = 10) were resistant to trimethoprim/sulfamethoxazole and rifampicin; 25.8% (n = 8) were resistant to erythromycin; 16.1% (n = 5) were resistant to teicoplanin; 6.5% (n = 2) were resistant to tetracycline. All Gram-positive bacteria were 100% susceptible to linezolid, gentamicin, tobramycin, levofloxacin, moxifloxacin, tigecycline, and nitrofurantoin. In antimicrobial susceptibility tests for the Gram-negative bacteria, 57.14% (n = 8) of the identified bacteria were resistant to ampicillin, whereas 50% (n = 7) were resistant to cefoxitin and ceftazidime. About 28.57% (n = 4) of the Gram-negative bacteria were resistant to ceftriaxone, trimethoprim/sulfamethoxazole. In addition, 21.43% (n = 3) were resistant to amoxicillin/clavulanic acid and cephalothin; 14.29% (n = 2) were resistant to cefepime and nitrofurantoin; 7.14% (n = 1) were resistant to piperacillin/tazobactam and tigecycline. However, all Gram-negative bacteria were susceptible to other examined antimicrobials. This is the first study that investigates the role of the hard tick as a potential reservoir for AMR pathogens within our region.

Seroprevalence of Toscana and sandfly fever Sicilian viruses in humans and livestock animals from western Saudi Arabia.

High seroprevalence rates of several phleboviruses have been reported in domestic animals and humans in sandfly-infested regions. Sandfly Fever Sicilian virus (SFSV) and Toscana virus (TOSV) are two of these viruses commonly transmitted by Phlebotomus sandflies. While SFSV can cause rapidly resolving mild febrile illness, TOSV could involve the central nervous system (CNS), causing diseases ranging from aseptic meningitis to meningoencephalitis. Sandfly-associated phleboviruses have not been investigated before in Saudi Arabia and are potential causes of infection given the prevalence of sandflies in the country. Here, we investigated the seroprevalence of SFSV and TOSV in the western region of Saudi Arabia in samples collected from blood donors, livestock animals, and animal handlers. An overall seroprevalence of 9.4% and 0.8% was found in humans for SFSV and TOSV, respectively. Seropositivity was significantly higher in non-Saudis compared to Saudis and increased significantly with age especially for SFSV. The highest seropositivity rate was among samples collected from animal handlers. Specifically, in blood donors, 6.4% and 0.7% tested positive for SFSV and TOSV nAbs, respectively. Animal handlers showed higher seroprevalence rates of 16% and 1% for anti-SFSV and anti-TOSV nAbs, respectively, suggesting that contact with livestock animals could be a risk factor. Indeed, sera from livestock animals showed seropositivity of 53.3% and 4.4% in cows, 27.5% and 7.8% in sheep, 2.2% and 0.0% in goats, and 10.0% and 2.3% in camels for SFSV and TOSV, respectively. Together, these results suggest that both SFSV and TOSV are circulating in the western region of Saudi Arabia in humans and livestock animals, albeit at different rates, and that age and contact with livestock animals could represent risk factors for infection with these viruses.

Molecular Characterization of Leishmania Species among Patients with Cutaneous Leishmaniasis in Asir Province, Saudi Arabia.

Anthroponotic cutaneous leishmaniais (ACL) and zoonotic cutaneous leishmaniasis (ZCL) caused by Leishmania tropica and Leishmania major, respectively, are endemic vector-borne diseases in southern Saudi Arabia. In 2021, an outbreak of cutaneous leishmaniasis occurred in the province of Asir. The main objective of our investigation was to analyze the epidemiological features of CL in southern Saudi Arabia. The ministry of health recorded 194 CL patients between January and December 2021 from the Asir province. Our findings showed that the majority of CL patients (87.1%) originated from the governorates of Khamis-Mushait and Abha. Most of the patients were males (62.3%). While CL affected all age groups, those under 13 years old were the most affected (38.1%). For both genders, CL patients were mostly Saudi citizens (90.7%) compared to non-Saudi expatriates. The majority of CL patients (75.2%) suffered from a single lesion, and the majority of lesions (61.3%) were located on the face. The seasonal prevalence of CL showed two peaks, a small one in July-August and a larger one in March. Of a total of 194 Giemsa slides samples, 188 showed positive amplification of Leishmania ITS1 gene. Based on PCR-RFLP and PCR-HMR, 183 patients showed positive amplification of L. tropica and five patients showed positive amplification of L. major. Phylogenetic analysis revealed a clear distinct separation between L. major and L. tropica sequences. Our results provided strong evidence of the pre-domination of L. tropica, the main etiological agent of ACL in Asir province. We reported for the first time the presence of L. major, an etiological agent of ZCL in the study areas. The co-circulation of ACL and ZCL highlighted the complexity of the epidemiology of CL in southern Saudi Arabia, and subsequently, further studies to identify competent vectors and reservoir hosts for the establishment of control strategies are needed.

Seroprevalence and Risk Factors Associated with Canine Leishmaniasis in Egypt.

BACKGROUND: Canine leishmaniasis (CanL) is caused by Leishmania infantum (L. infantum) that is transmitted by sand fly vectors with dogs acting as the main reservoir. METHODS: The present study aimed to determine the seroprevalence of CanL in dogs from Egypt and assessed the associated risk factors. The study was conducted from 2019 to 2020 in five governorates situated in Northern Egypt. Serum samples from 450 asymptomatic dogs were serologically examined by use of enzyme-linked immunosorbent assay (ELISA). RESULTS: Overall, the seroprevalence rate of CanL was 21.3% and the highest rates were observed in Cairo and Giza governorates. The univariable analysis revealed that the seropositivity of CanL was strongly related to the dogs' ages, length of hair, absence of veterinary care or application of insecticides, and the type of floor of their shelters. The risk factors that were found to be associated with CanL in exposed dogs were: age group 2-4 years old (OR = 12, 95% CI: 1.6-92.3); short hair (OR = 2.07, 95% CI: 1.2-3.6); absence of veterinary care (OR = 2.7, 95% CI: 1.3-5.8); no application of insecticides (OR = 3.09, 95% CI: 1.5-6.5) and their residence in a shelter with an earthen floor (OR = 1.42, 95% CI: 0.7-2.9). CONCLUSIONS: Based on the present results, CanL is present in Egyptian dogs and this increases the possibility of transmission by sand fly to humans with whom they have contact. Consequently, an efficient monitoring programme and effective control measures are important to reduce the risk of infection.

The emergence and transmission of COVID-19 in European countries, 2019-2020: a comprehensive review of timelines, cases and containment.

When it emerged in late 2019, COVID-19 was carried via travelers to Germany, France and Italy, where freedom of movement accelerated its transmission throughout Europe. However, effective non-pharmaceutical interventions introduced by European governments led to containment of the rapid increase in cases within European nations. Electronic searches were performed to obtain the number of confirmed cases, incident rates and non-pharmaceutical government measures for each European country. The spread and impact of non-pharmaceutical interventions throughout Europe were assessed and visualized. Specifically, heatmaps were used to represent the number of confirmed cases and incident rates for each of the countries over time. In addition, maps were created showing the number of confirmed cases and incident rates in Europe on three different dates (15 March, 15 April and 15 May 2020), which allowed us to assess the geographic and temporal patterns of the disease.

Chromosome-Scale Assembly of the Complete Genome Sequence of Porcisia hertigi, Isolate C119, Strain LV43.

Porcisia hertigi is a parasitic kinetoplastid first isolated from porcupines (Coendou rothschildi) in central Panama in 1965. We present the complete genome sequence of P. hertigi, isolate C119, strain LV43, sequenced using combined short- and long-read technologies. This complete genome sequence will contribute to our knowledge of the parasitic genus Porcisia.

Chromosome-Scale Assembly of the Complete Genome Sequence of Leishmania (Mundinia) martiniquensis, Isolate LSCM1, Strain LV760.

Leishmania (Mundinia) martiniquensis is a kinetoplastid parasite that was first isolated in 1995 on Martinique. We report the first complete genome for Leishmania martiniquensis from Asia, isolate LSCM1, strain LV760, which was sequenced using combined short-read and long-read technologies. This will facilitate greater understanding of the evolution of the geographically dispersed subgenus Mundinia.

Chromosome-Scale Assembly of the Complete Genome Sequence of Leishmania (Mundinia) orientalis, Isolate LSCM4, Strain LV768.

Leishmania (Mundinia) orientalis is a kinetoplastid parasite first isolated in 2014 in Thailand. We report the complete genome sequence of L. (M.) orientalis, sequenced using combined short-read and long-read technologies. This will facilitate greater understanding of this novel pathogen and its relationship to other members of the subgenus Mundinia.

Chromosome-scale genome sequencing, assembly and annotation of six genomes from subfamily Leishmaniinae.

We provide the raw and processed data produced during the genome sequencing of isolates from six species of parasites from the sub-family Leishmaniinae: Leishmania martiniquensis (Thailand), Leishmania orientalis (Thailand), Leishmania enriettii (Brazil), Leishmania sp. Ghana, Leishmania sp. Namibia and Porcisia hertigi (Panama). De novo assembly was performed using Nanopore long reads to construct chromosome backbone scaffolds. We then corrected erroneous base calling by mapping short Illumina paired-end reads onto the initial assembly. Data has been deposited at NCBI as follows: raw sequencing output in the Sequence Read Archive, finished genomes in GenBank, and ancillary data in BioSample and BioProject. Derived data such as quality scoring, SAM files, genome annotations and repeat sequence lists have been deposited in Lancaster University's electronic data archive with DOIs provided for each item. Our coding workflow has been deposited in GitHub and Zenodo repositories. This data constitutes a resource for the comparative genomics of parasites and for further applications in general and clinical parasitology.

LGAAP: Leishmaniinae Genome Assembly and Annotation Pipeline.

We present the LGAAP computational pipeline, which was successfully used to assemble six genomes of the parasite subfamily Leishmaniinae to chromosome-scale completeness from a combination of long- and short-read sequencing data. LGAAP is open source, and we suggest that it may easily be ported for assembly of any genome of comparable size (∼35 Mb).

Chromosome-Scale Assembly of the Complete Genome Sequence of Leishmania (Mundinia) sp. Ghana, Isolate GH5, Strain LV757.

Leishmania (Mundinia) sp. Ghana is a kinetoplastid parasite isolated in 2015 in Ghana. We report the complete genome sequence of L. (M.) sp. Ghana, sequenced using combined short-read and long-read technologies. This will facilitate greater understanding of this novel pathogen and its relationships within the subgenus Mundinia.

Chromosome-Scale Assembly of the Complete Genome Sequence of Leishmania (Mundinia) enriettii, Isolate CUR178, Strain LV763.

Leishmania (Mundinia) enriettii is a parasitic kinetoplastid first isolated from a guinea pig in Brazil in 1946. We present the complete genome sequence of L. (M.) enriettii, isolate CUR178, strain LV763, sequenced using combined short-read and long-read technologies. This will facilitate a greater understanding of the genome diversity within L. (M.) enriettii.

Reversed Immunoglycomics Identifies α-Galactosyl-Bearing Glycotopes Specific for Leishmania major Infection.

All healthy humans have high levels of natural anti-α-galactosyl (α-Gal) antibodies (elicited by yet uncharacterized glycotopes), which may play important roles in immunoglycomics: (a) potential protection against certain parasitic and viral zoonotic infections; (b) targeting of α-Gal-engineered cancer cells; (c) aiding in tissue repair; and (d) serving as adjuvants in α-Gal-based vaccines. Patients with certain protozoan infections have specific anti-α-Gal antibodies, elicited against parasite-derived α-Gal-bearing glycotopes. These glycotopes, however, remain elusive except for the well-characterized glycotope Galα1,3Galß1,4GlcNAcα, expressed by Trypanosoma cruzi. The discovery of new parasitic glycotopes is greatly hindered by the enormous structural diversity of cell-surface glycans and the technical challenges of classical immunoglycomics, a top-down approach from cultivated parasites to isolated glycans. Here, we demonstrate that reversed immunoglycomics, a bottom-up approach, can identify parasite species-specific α-Gal-bearing glycotopes by probing synthetic oligosaccharides on neoglycoproteins. This method was tested here seeking to identify as-yet unknown glycotopes specific for Leishmania major, the causative agent of Old-World cutaneous leishmaniasis (OWCL). Neoglycoproteins decorated with synthetic α-Gal-containing oligosaccharides derived from L. major glycoinositolphospholipids served as antigens in a chemiluminescent enzyme-linked immunosorbent assay using sera from OWCL patients and noninfected individuals. Receiver-operating characteristic analysis identified Galpα1,3Galfß and Galpα1,3Galfß1,3Manpα glycotopes as diagnostic biomarkers for L. major-caused OWCL, which can distinguish with 100% specificity from heterologous diseases and L. tropica-caused OWCL. These glycotopes could prove useful in the development of rapid α-Gal-based diagnostics and vaccines for OWCL. Furthermore, this method could help unravel cryptic α-Gal-glycotopes of other protozoan parasites and enterobacteria that elicit the natural human anti-α-Gal antibodies.

Cutaneous leishmaniasis and co-morbid major depressive disorder: A systematic review with burden estimates.

BACKGROUND: Major depressive disorder (MDD) associated with chronic neglected tropical diseases (NTDs) has been identified as a significant and overlooked contributor to overall disease burden. Cutaneous leishmaniasis (CL) is one of the most prevalent and stigmatising NTDs, with an incidence of around 1 million new cases of active CL infection annually. However, the characteristic residual scarring (inactive CL) following almost all cases of active CL has only recently been recognised as part of the CL disease spectrum due to its lasting psychosocial impact. METHODS AND FINDINGS: We performed a multi-language systematic review of the psychosocial impact of active and inactive CL. We estimated inactive CL (iCL) prevalence for the first time using reported WHO active CL (aCL) incidence data that were adjusted for life expectancy and underreporting. We then quantified the disability (YLD) burden of co-morbid MDD in CL using MDD disability weights at three severity levels. Overall, we identified 29 studies of CL psychological impact from 5 WHO regions, representing 11 of the 50 highest burden countries for CL. We conservatively calculated the disability burden of co-morbid MDD in CL to be 1.9 million YLDs, which equalled the overall (DALY) disease burden (assuming no excess mortality in depressed CL patients). Thus, upon inclusion of co-morbid MDD alone in both active and inactive CL, the DALY burden was seven times higher than the latest 2016 Global Burden of Disease study estimates, which notably omitted both psychological impact and inactive CL. CONCLUSIONS: Failure to include co-morbid MDD and the lasting sequelae of chronic NTDs, as exemplified by CL, leads to large underestimates of overall disease burden.

Old World cutaneous leishmaniasis treatment response varies depending on parasite species, geographical location and development of secondary infection.

BACKGROUND: In the Kingdom of Saudi Arabia (KSA), Leishmania major and L. tropica are the main causative agents of Old World cutaneous leishmaniasis (CL). The national CL treatment regimen consists of topical 1% clotrimazole/2% fusidic acid cream followed by 1-2 courses of intralesional sodium stibogluconate (SSG); however, treatment efficacy is highly variable and the reasons for this are not well understood. In this study, we present a complete epidemiological map of CL and determined the efficacy of the standard CL treatment regime in several endemic regions of KSA. RESULTS: Overall, three quarters of patients in all CL-endemic areas studied responded satisfactorily to the current treatment regime, with the remaining requiring only an extra course of SSG. The majority of unresponsive cases were infected with L. tropica. Furthermore, the development of secondary infections (SI) around or within the CL lesion significantly favoured the treatment response of L. major patients but had no effect on L. tropica cases. CONCLUSIONS: The response of CL patients to a national treatment protocol appears to depend on several factors, including Leishmania parasite species, geographical location and occurrences of SI. Our findings suggest there is a need to implement alternative CL treatment protocols based on these parameters.