Case Report: A Novel Virulent Clinical Isolate of Burkholderia pseudomallei Imported from Thailand Exhibiting Resistance to Trimethoprim-Sulfamethoxazole.

We present the investigation of a Burkholderia pseudomallei strain isolated from the urine of a 56-year-old male returning from a recreational trip to Thailand. The patient initially presented with fever and chills, and lobar as well as bladder and prostate involvement was demonstrated on imaging. Treatment first involved trimethoprim-sulfamethoxazole, the backbone of melioidosis therapy, but was discontinued upon the discovery of resistance to the drug through antimicrobial sensitivity testing via broth microdilution. Further analysis included genomic sequencing, immunotyping, and phenotypic assessment, including virulence testing in an animal model. Results indicated that this strain is a distinct, novel variant with antigenic similarity to other Thai strains displaying high virulence, with a significantly low murine intranasal lethal dose 50% inoculum. The emergence of such a strain, particularly if prevalent in tourist destinations in Thailand, could pose a substantial public health risk, highlighting the need for ongoing vigilance among infectious disease specialists and clinical microbiologists.

Highly sensitive extraction-free saliva-based molecular assay for rapid diagnosis of SARS-CoV-2.

The COVID-19 pandemic highlighted the necessity of fast, sensitive, and efficient methods to test large populations for respiratory viruses. The "gold standard" molecular assays for detecting respiratory viruses, such as quantitative polymerase chain reaction (qPCR) and reverse transcription qPCR (RT-qPCR), rely on invasive swab samples and require time-consuming and labor-intensive extraction processes. Moreover, the turnaround time for RT-qPCR-based assays is too lengthy for rapid screening. Extraction-free saliva-based methods provide a non-invasive sampling process with a fast turnaround time and are suitable for high-throughput applications. However, when used with a standard RT-qPCR system, the absence of extraction significantly reduces the assays' sensitivity. Here, using a novel optical modulation biosensing (OMB) platform, we developed a rapid and highly sensitive extraction-free saliva-based molecular assay. We blindly tested 364 paired nasopharyngeal swabs and saliva samples from suspected SARS-CoV-2 cases in Israel. Compared with the gold standard swab-based RT-qPCR assay, the sensitivity of the extraction-free saliva-based OMB assay is 90.7%, much higher than the sensitivity of extraction-free saliva-based RT-qPCR assay (77.8%) with similar specificity (95.3% and 97.6%, respectively). Moreover, out of 12 samples identified by the OMB-based assay as positive, 8 samples were collected from hospitalized patients in a COVID-19 ward and were verified to be SARS-CoV-2-positive upon admission, indicating that the actual clinical sensitivity and specificity of the OMB assay are higher. Considering its user-friendly saliva-based protocol, short and cost-effective extraction-free process, and high clinical accuracy, the OMB-based molecular assay is very suitable for high-throughput testing of large populations for respiratory viruses. IMPORTANCE: Three years after the SARS-CoV-2 outbreak, there are no molecular tests that combine low-cost and straightforward sample preparation, effective sample handling, minimal reagent and disposable requirements, high sensitivity, and high throughput required for mass screening. Existing rapid molecular techniques typically sacrifice certain requirements to meet others. Yet, localized outbreaks of novel viral diseases happen daily in different parts of the world. In this context, respiratory diseases are of specific importance, as they are frequently airborne and highly contagious, with the potential for a rapid global spread. The widely accepted opinion is that another pandemic is just a question of time. To ensure that the containment efforts for the upcoming "disease X" are successful, introducing rapid, high-throughput, and highly sensitive diagnostic methods for detecting and identifying pathogens is critical. A few months into the pandemic, saliva was suggested as a diagnostic matrix for SARS-CoV-2 detection. The collection of saliva does not require swabs and is minimally invasive. In particular, extraction-free saliva-based assays require fewer reagents and disposables, and therefore are faster and cheaper, offering an appealing alternative for low-income countries. Unfortunately, current extraction-free saliva-based detection methods, such as direct RT-qPCR or isothermal amplification, have either low sensitivity or low throughput. Therefore, we believe that the presented highly sensitive ht-OMBi platform and the extraction-free saliva-based molecular assay can become an essential tool in the infectious disease monitoring toolbox.

Overlooked monkeypox cases among men having sex with men during the 2022 outbreak - a retrospective study.

OBJECTIVES: The aim of this study was to characterize overlooked cases of patients with monkeypox infection in the 2022 outbreak. METHODS: Clinical characteristics of 26 patients who were misdiagnosed with other diseases were described. RESULTS: Of the 26 patients who were misdiagnosed, six (23%) were given a diagnosis of bacterial tonsillitis, six (23%) were diagnosed with primary syphilis, five (19.2%) with oral or genital herpes, and four (15.3%) with bacterial proctitis or anal abscess. The average time interval between missed and right diagnosis was 4.4 days. There was no difference in the missed cases between the early and the later month of the outbreak. CONCLUSION: Monkeypox might still be commonly overlooked, especially in patients presenting with fever and sore throat or solitary ulcer as sole manifestations.

Primary exposure to SARS-CoV-2 variants elicits convergent epitope specificities, immunoglobulin V gene usage and public B cell clones.

An important consequence of infection with a SARS-CoV-2 variant is protective humoral immunity against other variants. However, the basis for such cross-protection at the molecular level is incompletely understood. Here, we characterized the repertoire and epitope specificity of antibodies elicited by infection with the Beta, Gamma and WA1 ancestral variants and assessed their cross-reactivity to these and the more recent Delta and Omicron variants. We developed a method to obtain immunoglobulin sequences with concurrent rapid production and functional assessment of monoclonal antibodies from hundreds of single B cells sorted by flow cytometry. Infection with any variant elicited similar cross-binding antibody responses exhibiting a conserved hierarchy of epitope immunodominance. Furthermore, convergent V gene usage and similar public B cell clones were elicited regardless of infecting variant. These convergent responses despite antigenic variation may account for the continued efficacy of vaccines based on a single ancestral variant.

Unusual and Severe Complications of Acute Schistosomiasis in Travelers.

Acute schistosomiasis (ASC) is a hypersensitivity reaction seen mostly in nonimmune travelers and manifests mainly with fever, urticaria, and respiratory symptoms. We describe unusual severe presentations of ASC in 3 patients, including hip-monoarthritis, peri-myocarditis, and optic neuritis. In all 3 patients, clinical symptoms appeared or worsened after praziquantel administration.

The effect of ivermectin on the viral load and culture viability in early treatment of nonhospitalized patients with mild COVID-19 - a double-blind, randomized placebo-controlled trial.

OBJECTIVES: Ivermectin, an antiparasitic agent, also has antiviral properties. In this study, we aimed to assess whether ivermectin has anti-SARS-CoV-2 activity. METHODS: In this double-blinded trial, we compared patients receiving ivermectin for 3 days versus placebo in nonhospitalized adult patients with COVID-19. A reverse transcriptase-polymerase chain reaction from a nasopharyngeal swab was obtained at recruitment and every 2 days for at least 6 days. The primary endpoint was a reduction of viral load on the sixth day as reflected by cycle threshold level >30 (noninfectious level). The primary outcome was supported by the determination of viral-culture viability. RESULTS: Of 867 patients screened, 89 were ultimately evaluated per-protocol (47 ivermectin and 42 placeboes). On day 6, the odds ratio (OR) was 2.62 (95% confidence interval [CI]: 1.09-6.31) in the ivermectin arm, reaching the endpoint. In a multivariable logistic regression model, the odds of a negative test on day 6 were 2.28 times higher in the ivermectin group but reached significance only on day 8 (OR 3.70; 95% CI: 1.19-11.49, P = 0.02). Culture viability on days 2 to 6 was positive in 13.0% (3/23) of ivermectin samples versus 48.2% (14/29) in the placebo group (P = 0.008). CONCLUSION: There were lower viral loads and less viable cultures in the ivermectin group, which shows its anti-SARS-CoV-2 activity. It could reduce transmission in these patients and encourage further studies with this drug.

Primary exposure to SARS-CoV-2 variants elicits convergent epitope specificities, immunoglobulin V gene usage and public B cell clones.

An important consequence of infection with a SARS-CoV-2 variant is protective humoral immunity against other variants. The basis for such cross-protection at the molecular level is incompletely understood. Here we characterized the repertoire and epitope specificity of antibodies elicited by Beta, Gamma and ancestral variant infection and assessed their cross-reactivity to these and the more recent Delta and Omicron variants. We developed a high-throughput approach to obtain immunoglobulin sequences and produce monoclonal antibodies for functional assessment from single B cells. Infection with any variant elicited similar cross-binding antibody responses exhibiting a remarkably conserved hierarchy of epitope immunodominance. Furthermore, convergent V gene usage and similar public B cell clones were elicited regardless of infecting variant. These convergent responses despite antigenic variation may represent a general immunological principle that accounts for the continued efficacy of vaccines based on a single ancestral variant.

Existence of immunological memory response in true sero-negative individuals post COVID-19 molecular diagnosis.

Approximately 1-8% of individuals do not develop antibodies following SARS-CoV-2 infection (sero-negatives). One BNT162b2 dose resulted in potent humoral response in 14 sero-negatives and 15 sero-positives, significantly higher than the response of 15 naïve-individuals, to two doses suggesting that COVID-19 provoked a memory response in individuals without detectable antibodies.

Persistent abdominal symptoms in returning travellers: clinical and molecular findings.

BACKGROUND: Persistent abdominal symptoms (PAS) are the leading cause of post-travel morbidity although there is a paucity of evidence concerning the aetiology of this condition. Recently molecular methods for protozoa detection in stool have been introduced. Herein, we describe the clinical aspects and the prevalence of gastrointestinal protozoa in returning travellers with PAS. METHODS: From 2017 to 2019, clinical information and stool specimens from returning travellers with PAS were analysed for the presence of parasites using the Allplex-GI-Parasite-assay. Stool findings from symptomatic patients without a travel history were used as a comparator. RESULTS: During the 2-year study, 203 stool specimens from returning travellers were analysed. The median duration of symptoms before seeking care was 6 months, the most common symptoms were fatigue (79.2%), abdominal pain (75.7%) and loose stool (70.8%).Most of travellers had returned from Asia (57.6%), mainly from the Indian-subcontinent and only 52.6% were backpackers. Altogether, 36.9% samples were positive for protozoa, with Blastocystis hominis being the most common (26.6%) in samples, followed by Dientamoeba fragilis (18.7%), Giardia lamblia (3.0%) and Cryptosporidium spp (0.5%). The former two were dominant in all regions. In all cases but one, G. lamblia was acquired, but one were acquired in the Indian subcontinent (odds ratios 16.9; 95% confidence intervals: 1.9-148.3). Entamoeba histolytica was not detected. The demographic characterization of the 1359 non-travellers was comparable with the travellers. Among them D. fragilis was the most common followed by B. hominis, which was significantly less frequent compared among the travellers (16.7% vs 26.6%, P < 0.001). Average Cycle threshold values for each stool parasites were comparable between the two groups. CONCLUSION: Among returning travellers with PAS, more than one-third were positive for gastrointestinal protozoa. A low rate of giardia was found and no E. histolytica while B. hominis followed by D. fragilis were the dominant findings. Further studies are required to better understand the role of these protozoa in PAS.

Coronavirus disease 2019 (COVID-19) vaccination uptake among healthcare workers.

OBJECTIVE: To assess reasons for noncompliance with COVID-19 vaccination among healthcare workers (HCWs). DESIGN: Cohort observational and surveillance study. SETTING: Sheba Medical Center, a 1,600-bed tertiary-care medical center in Israel. PARTICIPANTS: The study included 10,888 HCWs including all employees, students, and volunteers. INTERVENTION: The BNT162b2 mRNA COVID-19 vaccine was offered to all HCWs of the hospital. Noncompliance was assessed, and pre-rollout and post-rollout surveys were conducted. Data regarding uptake of the vaccine as well as demographic data and compliance with prior influenza vaccination were collected, and 2 surveys were distributed. The survey before the rollout pertained to the intention to receive the vaccine, and the survey after the rollout pertained to all unvaccinated HCWs regarding causes of hesitancy. RESULTS: In the pre-rollout survey, 1,673 (47%) of 3,563 HCWs declared their intent to receive the vaccine. Overall, 8,108 (79%) HCWs received the COVID-19 vaccine within 40 days of rollout. In a multivariate logistic regression model, the factors that were significant predictors of vaccine uptake were male sex, age 40-59 years, occupation (paramedical professionals and doctors), high socioeconomic level, and compliance with flu vaccine. Among 425 unvaccinated HCWs who answered the second survey, the most common cause for hesitancy was the risk during pregnancy (31%). CONCLUSIONS: Although vaccine uptake among HCWs was higher than expected, relatively low uptake was observed among young women and those from lower socioeconomic levels and educational backgrounds. Concerns regarding vaccine safety during pregnancy were common and more data about vaccine safety, especially during pregnancy, might improve compliance.

Immunoglobulin (Ig)A seropositivity against SARS-CoV-2 in healthcare workers in Israel, 4 April to 13 July 2020: an observational study.

IntroductionThe COVID-19 pandemic has put healthcare workers (HCW) at significant risk. Presence of antibodies can confirm prior severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection.AimThis study investigates the prevalence of IgA and IgG antibodies against SARS-CoV-2 in HCW.MethodsPerformance of IgA and IgG antibody ELISA assays were initially evaluated in positive and negative SARS-CoV-2 serum samples. IgA and IgG antibodies against SARS-CoV-2 were measured in 428 asymptomatic HCW. We assessed the risk of two groups: HCW with high exposure risk outside work (HROW) residing in areas where COVID-19 was endemic (n = 162) and HCW with high exposure risk at work (HRAW) in a COVID-19 intensive care unit (ICU) (n = 97).ResultsSensitivities of 80% and 81.2% and specificities of 97.2% and 98% were observed for IgA and IgG antibodies, respectively. Of the 428 HCW, three were positive for IgG and 27 for IgA. Only 3/27 (11%) IgA-positive HCW had IgG antibodies compared with 50/62 (81%) in a group of previous SARS-CoV-2-PCR-positive individuals. Consecutive samples from IgA-positive HCW demonstrated IgA persistence 18-83 days in 12/20 samples and IgG seroconversion in 1/20 samples. IgA antibodies were present in 8.6% of HROW and 2% of HRAW.ConclusionsSARS-CoV-2 exposure may lead to asymptomatic transient IgA response without IgG seroconversion. The significance of these findings needs further study. Out of work exposure is a possible risk of SARS-CoV-2 infection in HCW and infection in HCW can be controlled if adequate protective equipment is implemented.

Anti-SARS-CoV-2 antibodies elicited by COVID-19 mRNA vaccine exhibit a unique glycosylation pattern.

Messenger RNA-based vaccines against COVID-19 induce a robust anti-SARS-CoV-2 antibody response with potent viral neutralization activity. Antibody effector functions are determined by their constant region subclasses and by their glycosylation patterns, but their role in vaccine efficacy is unclear. Moreover, whether vaccination induces antibodies similar to those in patients with COVID-19 remains unknown. We analyze BNT162b2 vaccine-induced IgG subclass distribution and Fc glycosylation patterns and their potential to drive effector function via Fcγ receptors and complement pathways. We identify unique and dynamic pro-inflammatory Fc compositions that are distinct from those in patients with COVID-19 and convalescents. Vaccine-induced anti-Spike IgG is characterized by distinct Fab- and Fc-mediated functions between different age groups and in comparison to antibodies generated during natural viral infection. These data highlight the heterogeneity of Fc responses to SARS-CoV-2 infection and vaccination and suggest that they support long-lasting protection differently.

A Serological Snapshot of COVID-19 Initial Stages in Israel by a 6-Plex Antigen Array.

The first case of SARS-CoV-2 was discovered in Israel in late February 2020. Three major outbreaks followed, resulting in over 800,000 cases and over 6,000 deaths by April 2021. Our aim was characterization of a serological snapshot of Israeli patients and healthy adults in the early months of the COVID-19 pandemic. Sera from 55 symptomatic COVID-19 patients and 146 healthy subjects (early-pandemic, reverse transcription-quantitative PCR [qRT-PCR]-negative), collected in Israel between March and April 2020, were screened for SARS-CoV-2-specific IgG, IgM, and IgA antibodies, using a 6-plex antigen microarray presenting the whole inactivated virus and five viral antigens: a stabilized version of the spike ectodomain (S2P), spike subunit 1 (S1), receptor-binding-domain (RBD), N-terminal-domain (NTD), and nucleocapsid (NC). COVID-19 patients, 4 to 40 days post symptom onset, presented specific IgG to all of the viral antigens (6/6) in 54 of the 55 samples (98% sensitivity). Specific IgM and IgA antibodies for all six antigens were detected in only 10% (5/55) and 4% (2/55) of the patients, respectively, suggesting that specific IgG is a superior serological marker for COVID-19. None of the qRT-PCR-negative sera reacted with all six viral antigens (100% specificity), and 48% (70/146) were negative throughout the panel. Our findings confirm a low seroprevalence of anti-SARS-CoV-2 antibodies in the Israeli adult population prior to the COVID-19 outbreak. We further suggest that the presence of low-level cross-reacting antibodies in naive individuals calls for a combined, multiantigen analysis for accurate discrimination between naive and exposed individuals. IMPORTANCE A 6-plex protein array presenting the whole inactivated virus and five nucleocapsid and spike-derived SARS-CoV-2 antigens was used to generate a serological snapshot of SARS-CoV-2 seroprevalence and seroconversion in Israel in the early months of the pandemic. Our findings confirm a very low seroprevalence of anti-SARS-CoV-2 antibodies in the Israeli adult population. We further propose that the presence of low-level nonspecific antibodies in naive individuals calls for a combined, multiantigen analysis for accurate discrimination between naive and exposed individuals enabling accurate determination of seroconversion. The developed assay is currently applied to evaluate immune responses to the Israeli vaccine during human phase I/II trials.

First reported nosocomial SARS-CoV-2 outbreak in a hospital-based laundry facility.

Nosocomial severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) outbreaks among health care workers have been scarcely reported so far. This report presents the results of an epidemiologic and molecular investigation of a SARS-CoV-2 outbreak among laundromat facility workers in a large tertiary centre in Israel. Following the first three reported cases of SARS-CoV-2 among laundromat workers, all 49 laundromat personnel were screened by qRT-PCR tests using naso- and oropharingeal swabs. Epidemiologic investigations included questionnaires, interviews and observations of the laundromat facility. Eleven viral RNA samples were then sequenced, and a phylogenetic analysis was performed using MEGAX.The integrated investigation defined three genetic clusters and helped identify the index cases and the assumed routes of transmission. It was then deduced that shared commute and public showers played a role in SARS-CoV-2 transmission in this outbreak, in addition to improper PPE use and social gatherings (such as social eating and drinking). In this study, we present an integrated epidemiologic and molecular investigation may help detect the routes of SARS-CoV-2 transmission, emphasising such routes that are less frequently discussed. Our work reinforces the notion that person-to-person transmission is more likely to cause infections than environmental contamination (e.g. from handling dirty laundry).

The role of mouthwash sampling in SARS-CoV-2 diagnosis.

BACKGROUND: The current practice of COVID-19 diagnosis worldwide is the use of oro-nasopharyngeal (ONP) swabs. Our study aim was to explore mouthwash (MW) as an alternative diagnostic method, in light of the disadvantages of ONP swabs. METHODS: COVID-19 outpatients molecular-confirmed by ONP swab were repeatedly examined with ONP swab and MW with normal saline (0.9%). Other types of fluids were compared to normal saline. The Cq values obtained with each method were compared. RESULTS: Among 137 pairs of ONP swabs and MW samples, 84.6% (116/137) of ONP swabs were positive by at least one of the genes (N, E, R). However MW detected 70.8% (97/137) of samples as positive, which means 83.6% (97/116) out of positive ONP swabs, missing mainly Cq value > 30. In both methods, the N gene was the most sensitive one. Therefore, MW samples targeting N gene, which was positive in 95/137 (69.3%), are comparable to ONP swabs targeting E and R genes which gave equal results-95/137 (69.3%) and 90/137 (65.7%), respectively. Comparing saline MW to distilled water gave equal results, while commercial mouth-rinsing solutions were less sensitive. CONCLUSIONS: MW with normal saline, especially when tested by N gene, can effectively detect COVID-19 patients. Furthermore, this method was not inferior when compared to R and E genes of ONP swabs, which are common targets in many laboratories around the world.

SARS-CoV-2 outbreak in a synagogue community: longevity and strength of anti-SARS-CoV-2 IgG responses.

Severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) pandemic is still ongoing along with the global vaccination efforts against it. Here, we aimed to understand the longevity and strength of anti-SARS-CoV-2 IgG responses in a small community (n = 283) six months following local SARS-COV-2 outbreak in March 2020. Three serological assays were compared and neutralisation capability was also determined. Overall 16.6% (47/283) of the participants were seropositive and 89.4% (42/47) of the IgG positives had neutralising antibodies. Most of the symptomatic individuals confirmed as polymerase chain reaction (PCR) positive during the outbreak were seropositive (30/32, 93.8%) and 33.3% of the individuals who quarantined with a PCR confirmed patient had antibodies. Serological assays comparison revealed that Architect (Abbott) targeting the N protein LIASON® (DiaSorin) targeting the S protein and enzyme-linked immunosorbent assay (ELISA) targeting receptor binding domain detected 9.5% (27/283), 17.3% (49/283) and 17% (48/283), respectively, as IgG positives. The latter two assays highly agreed (kappa = 0.89) between each other. In addition, 95%, (19/20, by ELISA) and 90.9% (20/22, with LIASON) and only 71.4% (15/21, by Architect) of individuals that were seropositive in May 2020 were found positive also in September. The unexpected low rate of overall immunity indicates the absence of un-noticed, asymptomatic infections. Lack of overall high correlation between the assays is attributed mainly to target-mediated antibody responses and suggests that using a single serological assay may be misleading.

Immunogenicity and safety of the BNT162b2 mRNA COVID-19 vaccine in people living with HIV-1.

OBJECTIVES: The immunogenicity and safety of the Pfizer-BioNTech BNT162b2 mRNA vaccine in people living with human immunodeficiency virus type 1 (PLWH) are unknown. We aimed to assess the immunogenicity and safety of this vaccine in PLWH. METHODS: In this prospective open study, we enrolled 143 PLWH, aged ≥18 years, who attended our clinic and 261 immunocompetent health-care workers (HCWs). Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) receptor binding domain (RBD) IgG and neutralizing antibodies were measured. Adverse events, viral load and CD4 cell counts were monitored. RESULTS: At a median of 18 days (interquartile range 14-21 days) after the second dose, anti-RBD-IgG was positive in 139/141 (98%) PLWH. Among HCWs, 258/261 (98.9%) developed anti-RBD-IgG at a median of 26 days (interquartile range 24-27 days) after the second dose. Following the second dose, immune sera neutralized SARS-CoV-2 pseudo-virus in 97% and 98% of PLWH and HCWs, respectively. Adverse events were reported in 60% of PLWH, mainly pain at the injection site, fatigue and headache. AIDS-related adverse events were not reported. Human immunodeficiency virus load increased in 3/143 (2%) patients from <40 copies/mL to ≤100 copies/mL. CD4+ T-cell count decreased from a geometric mean of 700 cells/µL (95% CI 648-757 cells/µL) to 633.8 cells/µL (95% CI 588-683 cells/µL) (p < 0.01). CONCLUSIONS: BNT162b2 mRNA vaccine appears immunogenic and safe in PLWH who are on antiretroviral therapy with unsuppressed CD4 count and suppressed viral load.

Establishing a COVID-19 treatment centre in Israel at the initial stage of the outbreak: challenges, responses and lessons learned.

Anticipating the need for a COVID-19 treatment centre in Israel, a designated facility was established at Sheba Medical Center-a quaternary referral centre. The goals were diagnosis and treatment of patients with COVID-19 while protecting patients and staff from infection and ensuring operational continuity and treatment of patients with non-COVID. Options considered included adaptation of existing wards, building a tented facility and converting a non-medical structure. The option chosen was a non-medical structure converted to a hospitalisation facility suited for COVID-19 with appropriate logistic and organisational adaptations. Operational principles included patient isolation, unidirectional workflow from clean to contaminated zones and minimising direct contact between patients and caregivers using personal protection equipment (PPE) and a multimodal telemedicine system. The ED was modified to enable triage and treatment of patients with COVID-19 while maintaining a COVID-19-free environment in the main campus. This system enabled treatment of patients with COVID-19 while maintaining staff safety and conserving the operational continuity and the ability to continue delivery of treatment to patients with non-COVID-19.