A Longitudinal Study in Tunisia to Assess the Anti-RBD IgG and IgA Responses Induced by Three Different COVID-19 Vaccine Platforms.

BACKGROUND: Vaccination constitutes the best strategy against COVID-19. In Tunisia, seven vaccines standing for the three main platforms, namely RNA, viral vector, and inactivated vaccines, have been used to vaccinate the population at a large scale. This study aimed to assess, in our setting, the kinetics of vaccine-induced anti-RBD IgG and IgA antibody responses. METHODS: Using in-house developed and validated ELISA assays, we measured anti-RBD IgG and IgA serum antibodies in 186 vaccinated workers at the Institut Pasteur de Tunis over 12 months. RESULTS: We showed that RNA vaccines were the most immunogenic vaccines, as compared to alum-adjuvanted inactivated and viral-vector vaccines, either in SARS-CoV-2-naïve or in SARS-CoV-2-experienced individuals. In addition to the IgG antibodies, the vaccination elicited RBD-specific IgAs. Vaccinated individuals with prior SARS-CoV-2 infection exhibited more robust IgG and IgA antibody responses, as compared to SARS-CoV-2-naïve individuals. CONCLUSIONS: After following up for 12 months post-immunization, we concluded that the hierarchy between the platforms for anti-RBD antibody-titer dynamics was RNA vaccines, followed by viral-vector and alum-adjuvanted inactivated vaccines.

Immunogenicity of Mix-and-Match CoronaVac/BNT162b2 Regimen versus Homologous CoronaVac/CoronaVac Vaccination: A Single-Blinded, Randomized, Parallel Group Superiority Trial.

(1) Background: This study aimed to compare the immunogenicity of the mix-and-match CoronaVac/BNT162b2 vaccination to the homologous CoronaVac/CoronaVac regimen. (2) Methods: We conducted a simple-blinded randomized superiority trial to measure SARS-CoV-2 neutralization antibodies and anti-spike receptor binding domain (RBD) IgG concentrations in blood samples of participants who had received the first dose of CoronaVac vaccine followed by a dose of BNT162b2 or CoronaVac vaccine. The primary endpoint for immunogenicity was the serum-neutralizing antibody level with a percentage of inhibition at 90% at 21-35 days after the boost. A difference of 25% between groups was considered clinically relevant. (3) Results: Among the 240 eligible participants, the primary endpoint data were available for 100 participants randomly allocated to the mix-and-match group versus 99 participants randomly allocated to the homologous dose group. The mix-and-match regimen elicited significantly higher levels of neutralizing antibodies (median level of 96%, interquartile range (IQR) (95-97) versus median level of 94%, IQR (81-96) and anti-spike IgG antibodies (median level of 13,460, IQR (2557-29,930) versus median level of 1190, IQR (347-4964) compared to the homologous group. Accordingly, the percentage of subjects with a percentage of neutralizing antibodies > 90% was significantly higher in the mix-and-match group (90.0%) versus the homologous (60.6%). Interestingly, no severe events were reported within 30 days after the second dose of vaccination in both groups. (4) Conclusions: Our data showed the superiority of the mix-and-match CoronaVac/BNT162b2 vaccination compared to the CoronaVac/CoronaVac regimen in terms of immunogenicity, thus constituting a proof-of-concept study supporting the use of inactivated vaccines in a mix-and-match strategy while ensuring good immunogenicity and safety.

Incidence and risk factors of SARS-CoV-2 infection among workers in a public health laboratory in Tunisia.

The aim of this study was to measure the extent of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection among workers at the Institut Pasteur de Tunis (IPT), a public health laboratory involved in the management of the COVID-19 pandemic in Tunisia, and to identify risk factors for infection in this occupational setting. A cross-sectional survey was conducted on IPT workers not vaccinated against coronavirus disease 2019 (COVID-19). Participants completed a questionnaire that included a history of reverse transcription-polymerase chain reaction (RT-PCR)-confirmed SARS-CoV-2 infection. Immunoglobulin G antibodies against the receptor-binding domain of the spike antigen (anti-S-RBD IgG) and the nucleocapsid protein (anti-N IgG) of the SARS-CoV-2 virus were detected by enzyme-linked immunoassay (ELISA). A multivariate analysis was used to identify factors significantly associated with SARS-CoV-2 infection. A total of 428 workers were enrolled in the study. The prevalence of anti-S-RBD and/or anti-N IgG antibodies was 32.9% [28.7-37.4]. The cumulative incidence of SARS-CoV-2 infection (positive serology and/or previous positive RT-PCR test) was 40.0% [35.5-44.9], while the proportion with asymptomatic infection was 32.9%. One-third of the participants with RT-PCR-confirmed infection tested seronegative more than 90 days postinfection. Participants aged over 40 and laborers were more susceptible to infection (adjusted OR [AOR] = 1.65 [1.08-2.51] and AOR = 2.67 [1.45-4.89], respectively), while tobacco smokers had a lower risk of infection (AOR = 0.54 [0.29-0.97]). The SARS-CoV-2 infection rate among IPT workers was not significantly different from that detected concurrently in the general population. Hence, the professional activities conducted in this public health laboratory did not generate additional risk to that incurred outside the institute in day-to-day activities.

Healed Lesions of Human Cutaneous Leishmaniasis Caused By Leishmania major Do Not Shelter Persistent Residual Parasites.

In human cutaneous leishmaniasis (HCL) caused by Leishmania (L.) major, the cutaneous lesions heal spontaneously and induce a Th1-type immunity that confers solid protection against reinfection. The same holds true for the experimental leishmaniasis induced by L. major in C57BL/6 mice where residual parasites persist after spontaneous clinical cure and induce sustainable memory immune responses and resistance to reinfection. Whether residual parasites also persist in scars of cured HCL caused by L. major is still unknown. Cutaneous scars from 53 volunteers with healed HCL caused by L. major were biopsied and the tissue sample homogenates were analyzed for residual parasites by four methods: i) microscope detection of amastigotes, ii) parasite culture by inoculation on biphasic medium, iii) inoculation of tissue exctracts to the footpad of BALB/c mice, an inbred strain highly susceptible to L. major, and iv) amplification of parasite kDNA by a highly sensitive real-time PCR (RT-PCR). Our results show that the scars of healed lesions of HCL caused by L. major do not contain detectable residual parasites, suggesting that this form likely induces a sterile cure at least within the scars. This feature contrasts with other Leishmania species causing chronic, diffuse, or recidivating forms of leishmaniasis where parasites do persist in healed lesions. The possibility that alternative mechanisms to parasite persistence are needed to boost and maintain long-term immunity to L. major, should be taken into consideration in vaccine development against L. major infection.

Knowledge attitudes and practices toward seasonal influenza vaccine among pregnant women during the 2018/2019 influenza season in Tunisia.

BACKGROUND: The uptake and acceptance of the influenza vaccine (IV) among pregnant women remain unknown in Tunisia despite the increased influenza-related complications and death. The present study aimed to assess the IV uptake and acceptability and to describe related knowledge and attitudes among pregnant women in Tunisia. METHODS: A cross-sectional study was conducted in 84 Tunisian healthcare facilities over a period of three months (from March to May 2019). All pregnant women aged ≥18 years who sought antenatal care in related health structures were included in this study based on a multistage self-weighted sampling. We measured knowledge and attitudes towards the IV and assessed factors related to willingness for its uptake. RESULTS: The questionnaire was completed by 1157 pregnant women. More than half of the participants (60.2%; 95% confidence interval [CI] [57.3%-63.0%]) reported awareness about the IV. Among included PW, only 4.6%; 95% CI [3.5%-6.1%] received it during their current pregnancy. However, (36.8%; 95% CI [34.0%-39.6%]) declared their willingness to receive the vaccine in the next pregnancy. Recommendation by healthcare providers, identified to be the most trustful source of information, was the main reason for acceptance. However, the intention to accept the IV by pregnant women was significantly associated with such recommendation and perceived safety and effectiveness of this vaccine. CONCLUSION: Antenatal care visits are a precious opportunity that should not be missed by health care providers and especially gynecologists to promote the IV uptake by pregnant women in Tunisia.

Intra-Specific Diversity of Leishmania major Isolates: A Key Determinant of Tunisian Zoonotic Cutaneous Leishmaniasis Clinical Polymorphism.

The clinical expression of zoonotic cutaneous leishmaniasis (ZCL) caused by Leishmania (L.) major parasites has a broad spectrum ranging from asymptomatic infection to self-limited cutaneous sores or severe disease. In concert with the host immune responses, the vector variability and the number of bites, genetic variation between L. major isolates might impact on the clinical output of the disease. We investigated herein the intra-specific variability of L. major field isolates independently of host or vector factors and then tried to correlate parasite variability to ZCL severity in corresponding patients. Several assays were applied, i.e., in vivo pathogenicity of promastigotes in a BALB/c mice model, resistance/sensibility to complement lysis, in vitro growth kinetics, and expression of different lectins on the promastigote surface. Combining all these parameters allowed us to conclude that the resistance to complement lysis and PNA/Jacalin lectins binding to parasite surfaces are important markers of parasite virulence. These factors correlate significantly with clinic polymorphism of ZCL and modestly with genetic micro-heterogeneity, a characteristic we previously revealed with a MLMT profile.

Influenza vaccination knowledge, attitudes, and practices among Tunisian elderly with chronic diseases.

BACKGROUND: Generally, seasonal influenza does not cause severe infection in healthy adults, but for the elderly, an infection can pose a serious health concern. Although several measures can help prevent influenza, vaccination is considered the most effective. This study aimed to assess influenza vaccine uptake among elderly with chronic diseases in Tunisia during the 2018-2019 influenza season, and to identify knowledge, attitudes and barriers associated with influenza vaccine uptake. METHODS: During influenza season of 2018-2019, we conducted a national cross-sectional study among elderly with chronic disease who were attending primary and secondary health care facilities in Tunisia. We collected data regarding practices, general knowledge and attitudes related to influenza and influenza vaccine, using a standardized questionnaire. A multivariate analysis by logistic regression was performed to assess the factors influencing willingness to receive influenza vaccine. RESULTS: Among the 1191 surveyed elderly, 19.4% (95%CI 14.1-21.9) were vaccinated during the 2018-2019 influenza season and 64.7% (61.9-67.3) expressed willingness to be vaccinated in the next season regardless of vaccination status in the 2018-2019 season. Previous vaccination in the 2018-2019 influenza season was the most significantly associated factor with willingness to receive influenza vaccine (adjusted OR = 16.5 [3.7-72.4]). Significant associations were also observed between knowledge of influenza severity for the elderly as well as for those with chronic diseases and willingness to be vaccinated (p < 0.01). Likewise, participants who were convinced by flu vaccine effectiveness and those who were not concerned about vaccine side effects were more likely to be vaccinated (p < 0.001). The main reason that may lead to vaccine acceptance was a doctor's recommendation (41.1%), while the two main reasons that may lead to vaccine refusal were concerns about side effects (71.5%) and a belief that vaccine was ineffective in averting influenza illness (33.9%). Doctors were the most trusted source for information about influenza vaccine (91.5%). CONCLUSION: Our study revealed low influenza vaccination coverage among Tunisian elderly with chronic diseases believed to be at higher risk for severe acute respiratory infections and death if infected with influenza. Treating physicians' role in promoting influenza vaccination in this high-risk group seems to be crucial.

Knowledge, attitudes and uptake related to influenza vaccine among healthcare workers during the 2018-2019 influenza season in Tunisia.

BACKGROUND: The influenza vaccine (IV) is considered the most effective strategy to prevent seasonal influenza infection and annual vaccination of healthcare workers (HCWs) is recommended by the World Health Organization given their high mixing with patients. We assessed IV uptake among HCWs in the 2018-2019 season and explored their knowledge and attitudes regarding influenza immunization. METHODS: A cross-sectional study was conducted in 150 representative Tunisian health facilities from March to May 2019. We recruited 1231 HCWs with direct patient contact using self-weighted multistage sampling. Univariate and multivariate logistic regression analyses permitted to assess the factors associated with IV uptake in the 2018-2019 influenza season. RESULTS: Among 1231 health professionals enrolled in this study, less than half (36.6, 95% confidence interval [CI]: 33.9-39.4) received the IV at least once in their lives and only 15.3% (CI: 13.3-17.4) were vaccinated against influenza in the 2018-2019 influenza season. High confidence regarding IV efficacy, belief about the mandatory character of influenza vaccination for HCWs, and IV uptake in the 4 years preceding the 2018-2019 influenza season were independently associated with higher IV uptake by multivariate analysis. However, participants with high educational level were less likely to receive the IV than those with the lowest educational level. CONCLUSIONS: Our study revealed a low vaccination rate among Tunisian HCWs confirming the importance of tailored education programs targeting this population.

Psycho-social impacts, experiences and perspectives of patients with Cutaneous Leishmaniasis regarding treatment options and case management: An exploratory qualitative study in Tunisia.

Although non-fatal and mostly self-healing in the case of Leishmania (L.) major, cutaneous leishmaniasis (CL) is mainly treated to reduce lesion healing time. Less attention is paid to the improvement of scars, especially in aesthetically relevant areas of the body, which can dramatically affect patients' wellbeing. We explored patients' perspectives about treatment options and the social and psychological burden of disease (lesion and scar). Individual in-depth interviews were conducted with ten confirmed CL patients at two L. major endemic sites in Southern Tunisia (Sidi Bouzid and Gafsa). Participants were selected using a sampling approach along a spectrum covering e.g. age, sex, and clinical presentation. Patients' experiences, opinions and preferences were explored, and their detailed accounts gave an insight on the impact of CL on their everyday lives. The impact of CL was found to be considerable. Most patients were not satisfied with treatment performance and case management. They expected a shorter healing time and better accessibility of the health system. Tolerance of the burden of disease was variable and ranged from acceptance of hidden scars to suicidal thoughts resulting from the fear to become handicapped, and the stress caused by close relatives. Some believed CL to be a form of skin cancer. Unexpectedly, this finding shows the big gap between the perspectives of patients and assumptions of health professionals regarding this disease. This study provided valuable information for better case management emphasizing the importance of improving communication with patients, and accessibility to treatment. It generated context-specific knowledge to policy makers in Tunisia to implement effective case management in a country where access to treatment remains a challenge due to socio-economic and geographic barriers despite a long tradition in CL control.

A prospective cohort study of Cutaneous Leishmaniasis due to Leishmania major: Dynamics of the Leishmanin skin test and its predictive value for protection against infection and disease.

BACKGROUND: Leishmanin Skin Test (LST) is considered as a useful indicator of past infection by Leishmania parasites. However, the temporal dynamics of a positive LST under different epidemiologic scenarios and whether it relates to the protection against the recurrence of an overt disease are not fully documented. METHODOLOGY/PRINCIPAL FINDINGS: We report here on a population based prospective study conducted on 2686 individuals living in two foci located in Central Tunisia, to assess over a one-year epidemiologic season, the incidence of Leishmania (L.) major infection and disease and changes in LST reactivity. The two foci were both endemic for Cutaneous Leishmaniasis (CL) due to L. major, but contrasted in their history for this disease (ie: an old focus versus a recent focus). We found that most infections occurred in the new focus (290/1000; 95% CI: 265-315 person-years) with an incidence rate of CL lesions 2.4 times higher than in the old focus. Likewise, the rates of LST reactivity reversion and loss, in the new focus, were 99/1000[38-116] person-years and 14/1000[8-21] person-years, respectively. Loss of LST reactivity was not noticed in the old focus. Interestingly, the incidence rates of symptomatic infection did not differ significantly according to the LST status at enrolment (negative versus positive) between the combined foci and the new one. CONCLUSIONS/SIGNIFICANCE: Our findings confirm LST as a good tool for assessing L. major cryptic infection. However, the instability of the LST positivity in new foci should be considered as an important confounder of the outcome of this infection when developing a research protocol for vaccine trial.

Hepatitis C virus epidemiology in Central-West Tunisia: a population-based cross-sectional study.

This study aimed to assess the seroprevalence, viraemia and genotype distribution of hepatitis C virus (HCV) in a region in Central-West Tunisia. A door-to-door cross-sectional study was conducted on a randomly selected sample. A total of 3178 individuals aged 5 to 74 years and members of 935 families were investigated. Seroprevalence of HCV was assessed using ELISA tests. The viral load was determined by real-time RT-PCR, and HCV genotyping was conducted by amplification and sequencing in the NS5b genomic region. The global prevalence of HCV antibodies was 3.32% (95% confidence interval [CI]: 2.72-4.00). It was significantly higher in women: 4.47% vs. 2.16% in men, p = 0.001. Seroprevalence increased with age, and the highest rates were found in the 50- to 59-year-old age group (12.90%, 95% CI: 9.45-16.86), suggesting a cohort effect with very low contribution of intrafamilial transmission. Genotyping showed a predominance of subtype 1b (84.6%), with cocirculation of subtypes 2c (9.6%), 1a (1.9%), 1d (1.9%) and 2k (1.9%), similar to the previously reported genotype distribution in Tunisia and with no genetic clusters specific to the study region. These results indicate a higher endemicity of HCV infection when compared to the previously reported nationwide surveillance data. This study provides valuable data that can contribute to current strategies to eliminate hepatitis C.

First Report on Natural Infection of Phlebotomus sergenti with Leishmania tropica in a Classical Focus of Leishmania major in Tunisia.

In Tunisia, chronic cutaneous leishmaniasis due to Leishmania tropica is an important health problem. Its spreading has not been fully elucidated. Information on sandfly vectors, as well as their associated Leishmania species, is of paramount importance since vector dispersion is one of the major factors responsible for pathogen dissemination. Ninety-seven unfed females belonging to the genera Sergentomyia and Phlebotomus were collected between June and August 2015 using sticky paper traps. Polymerase chain reaction-restriction fragment length polymorphism analysis of the internal transcribed spacer 1and sequencing were used for Leishmania detection and identification. In total, 650 sandflies were captured and identified (380 males and 270 females). Ninety-seven unfed females were tested for the presence of Leishmania parasite DNA. Six Phlebotomus sergenti were found positive for L. tropica. This novel finding enhances the understanding of the cycle extension of L. tropica outside its original focus of Tataouine.

Insight into the global evolution of Rodentia associated Morbilli-related paramyxoviruses.

One portion of the family Paramyxoviridae is a group of Unclassified Morbilli-Related Viruses (UMRV) recently recognized in wild small mammals. At a global level, the evolutionary history of these viruses is not properly understood and the relationships between UMRV and their hosts still remain largely unstudied. The present study revealed, for the first time, that Rodentia associated UMRV emerged from a common ancestor in southern Africa more than 4000 years ago. Sequenced UMRV originating from different regions in the world, clustered into four well-supported viral lineages, which suggest that strain diversification occurred during host dispersal and associated exchanges, with purifying selection pressure as the principal evolutionary force. In addition, multi-introductions on different continents and islands of Rodentia associated UMRV and spillover between rodent species, most probably Rattus rattus, were detected and indicate that these animals are implicated in the vectoring and in the worldwide emergence of this virus group. The natural history and the evolution dynamics of these zoonotic viruses, originating from and hosted by wild animals, are most likely shaped by commensalism related to human activities.

Involvement of different CD4(+) T cell subsets producing granzyme B in the immune response to Leishmania major antigens.

The nature of effector cells and the potential immunogenicity of Leishmania major excreted/secreted proteins (LmES) were evaluated using peripheral blood mononuclear cells (PBMCs) from healed zoonotic cutaneous leishmaniasis individuals (HZCL) and healthy controls (HC). First, we found that PBMCs from HZCL individuals proliferate and produce high levels of IFN-γ and granzyme B (GrB), used as a marker of activated cytotoxic T cells, in response to the parasite antigens. IFN-γ is produced by CD4(+) T cells, but unexpectedly GrB is also produced by CD4(+) T cells in response to stimulation with LmES, which were found to be as effective as soluble Leishmania antigens to induce proliferation and cytokine production by PBMCs from immune individuals. To address the question of regulatory T cell (Tregs) involvement, the frequency of circulating Tregs was assessed and found to be higher in HZCL individuals compared to that of HC. Furthermore, both CD4(+)CD25(+) and CD4(+)CD25(-) T cells, purified from HZCL individuals, produced IFN-γ and GrB when stimulated with LmES. Additional experiments showed that CD4(+)CD25(+)CD127(dim/-) Tregs were involved in GrB production. Collectively, our data indicate that LmES are immunogenic in humans and emphasize the involvement of CD4(+) T cells including activated and regulatory T cells in the immune response against parasite antigens.

Prevalence and determinants of Leishmania major infection in emerging and old foci in Tunisia.

BACKGROUND: Zoonotic Cutaneous Leishmaniasis (ZCL) due to Leishmania major (L. major) is still a serious public health problem in Tunisia. This study aimed to compare the prevalence and risk factors associated with L. major infection in old and new foci using leishmanin skin test (LST) in central Tunisia. METHODS: A cross sectional household survey was carried out between January and May 2009 on a sample of 2686 healthy individuals aged between 5 and 65 years. We determined the prevalence of L. major infection using the LST. Risk factors of LST positivity were assessed using a logistic regression model. RESULTS: The overall prevalence of LST positivity was 57% (95% CI: 53-59). The prevalence of L. major infection was significantly higher in the old focus (99%; 95% CI: 98-100) than in the emerging foci (43%; 95% CI: 39-46) (p = <0.001). Multivariate analysis of LST positivity risk factors showed that age, the nature of the foci (old/emerging), personal and family history of ZCL are determinants of positive LST results. CONCLUSION: The results updated the current epidemiologic profile of ZLC in central Tunisia. Past history of transmission in a population should be considered as a potential confounder in future clinical trials for drugs and vaccines against L. major cutaneous leishmaniasis.

Topical paromomycin with or without gentamicin for cutaneous leishmaniasis.

BACKGROUND: There is a need for a simple and efficacious treatment for cutaneous leishmaniasis with an acceptable side-effect profile. METHODS: We conducted a randomized, vehicle-controlled phase 3 trial of topical treatments containing 15% paromomycin, with and without 0.5% gentamicin, for cutaneous leishmaniasis caused by Leishmania major in Tunisia. We randomly assigned 375 patients with one to five ulcerative lesions from cutaneous leishmaniasis to receive a cream containing 15% paromomycin-0.5% gentamicin (called WR 279,396), 15% paromomycin alone, or vehicle control (with the same base as the other two creams but containing neither paromomycin nor gentamicin). Each lesion was treated once daily for 20 days. The primary end point was the cure of the index lesion. Cure was defined as at least 50% reduction in the size of the index lesion by 42 days, complete reepithelialization by 98 days, and absence of relapse by the end of the trial (168 days). Any withdrawal from the trial was considered a treatment failure. RESULTS: The rate of cure of the index lesion was 81% (95% confidence interval [CI], 73 to 87) for paromomycin-gentamicin, 82% (95% CI, 74 to 87) for paromomycin alone, and 58% (95% CI, 50 to 67) for vehicle control (P<0.001 for each treatment group vs. the vehicle-control group). Cure of the index lesion was accompanied by cure of all other lesions except in five patients, one in each of the paromomycin groups and three in the vehicle-control group. Mild-to-moderate application-site reactions were more frequent in the paromomycin groups than in the vehicle-control group. CONCLUSIONS: This trial provides evidence of the efficacy of paromomycin-gentamicin and paromomycin alone for ulcerative L. major disease. (Funded by the Department of the Army; ClinicalTrials.gov number, NCT00606580.).

Heterogeneity of hepatitis B transmission in Tunisia: risk factors for infection and chronic carriage before the introduction of a universal vaccine program.

UNLABELLED: A population-based sero-epidemiological study enrolled 9486 volunteers in two governorates, Béja in the north and Tataouine in the south of Tunisia, in order to assess the magnitude of HBV transmission heterogeneity between the north and the south and within the same governorate, as well as the risk factors associated with infection and chronic carriage. RESULTS: The overall prevalence of anti-HBc, HBsAg and chronic carriage was 28.5, 5.3 and 2.9%, respectively. Significant differences were observed between the two governorates according to anti-HBc (32.1% in Béja and 27.8% in Tataouine; p=0.005) and HBsAg prevalence (4.2% in Béja and 5.6% in Tataouine; p=0.001). Significant differences were noticed between districts revealing important heterogeneity in HBV transmission within the same governorate (HBsAg ranged from 12 to <2% within the same governorate). At the individual level, the presence of a family member infected with HBV, scarification practices, needle practices in the Primary Care Center and gender (male) significantly increased the risk of anti-Hbc, HBsAg positivity and chronic carriage of infection while existence of sanitation in the house was found to be protective. The basic reproductive number and the force of infection confirmed the heterogeneity of transmission. Horizontal transmission within the family explains hyperendemic clusters in Tunisia.