Corrigendum: Exploring genetic variability of Giardia duodenalis and Enterocytozoon bieneusi in raw vegetables and fruits: implications for food safety and public health in Mozambique.

[This corrects the article DOI: 10.3389/fmicb.2023.1223151.].

Exploring genetic variability of Giardia duodenalis and Enterocytozoon bieneusi in raw vegetables and fruits: implications for food safety and public health in Mozambique.

Giardia duodenalis and Enterocytozoon bieneusi are etiological agents of enteric diseases characterized by diarrhea that can progress to chronicity in humans, especially in children and in immunocompromised patients. This study aims to assess the genetic pattern of G. duodenalis and E. bieneusi detected in vegetables and fruits commercialized in Maputo markets, Mozambique and determine their public health importance. Eight study points were sampled: a farmer zone, a wholesale, four retail markets, and two supermarkets in Maputo city, where eight types of horticultural products were purchased. Using nested-PCR methods, 2.8% (9/321) and 1.3% (4/321) of samples monitored were positive for G. duodenalis and E. bieneusi, respectively. Based on the analysis of the ß-giardin and ITS rRNA sequences of G. duodenalis and E. bieneusi detected, respectively, four different sequences of G. duodenalis (three novel sequences: BgMZ1, BgMZ2, and BgMZ3, and one known sequence) all from assemblage B and three genotypes of E. bieneusi (two novel sequences: EbMZ4 and EbMZ5, and one known sequence: KIN-1) from group 1. These microorganisms were found and characterized for the first time in horticultural products in Maputo markets. All identified G. duodenalis and E. bieneusi display high genetic similarity within their ß-giardin and ITS rRNA sequences, respectively, having been clustered into assemblages and genotypes with high zoonotic transmission potential. Our study may represent a relevant step in the understanding of these intestinal pathogens in association with fresh vegetables and fruits for human consumption, for a better and broader "One Health" approach.

Intestinal Parasites in Children up to 14 Years Old Hospitalized with Diarrhea in Mozambique, 2014-2019.

Diarrhea remains a public health problem in Mozambique, even with control strategies being implemented. This analysis aimed to determine the proportion and factors associated with intestinal parasitic infection (IPI) in children up to 14 years old with diarrheal disease, in the southern, central and northern regions of Mozambique. A single diarrheal sample of 1424 children was collected in hospitals and examined using the formol-ether concentration and modified Ziehl−Neelsen techniques to identify intestinal parasites using optical microscopy. Sociodemographic characteristics were obtained by questionnaires. Descriptive statistics and cross-tabulation were performed, and p-values <0.05 were considered statistically significant. A single IPI was detected in 19.2% (273/1424) of the children. Cryptosporidium spp. was the most common parasite (8.1%; 115/1424). Polyparasitism was seen in 26.0% (71/273), with the co-infection of Ascaris lumbricoides and Trichuris trichiura (26.8%; 19/71) being the most common. Age and province were related to IPI (p-value < 0.05). The highest occurrence of IPI was observed in the wet period (October to March), with 21.9% (140/640), compared to the dry period (April to September), with 16.9% (131/776) (p-value = 0.017). Cryptosporidium spp. and the combination of A. lumbricoides/T. trichiura were the main intestinal parasites observed in children hospitalized with diarrhea in Mozambique.

Occurrence of Intestinal Parasites of Public Health Significance in Fresh Horticultural Products Sold in Maputo Markets and Supermarkets, Mozambique.

(1) Background: Giardia duodenalis and Cryptosporidium are important neglected parasites associated with diarrhea, such as the emerging Enterocytozoon bieneusi. All three are foodborne parasites raising concerns in public health. This study intended to understand the intestinal parasite occurrence with emphasis on G. duodenalis, Cryptosporidium, and E. bieneusi in fresh fruits/vegetables sold in the main municipal markets of Maputo city, Mozambique. (2) Methods: A total of 321 fresh horticultural products were purchased in the rainy and dry seasons (five markets/two supermarkets/one agricultural zone). Light microscopy (LM) and PCR analysis were performed. (3) Results: By LM and/or PCR, 29.3% of the samples presented at least one parasite (rainy season: 22.9%; dry season: 35.1%). The most contaminated horticultural products: collected in dry season, from Zimpeto and Fajardo markets, lettuce and pointed white cabbage. Overall, 3.7% of G. duodenalis, 1.3% of E. bieneusi, and other intestinal parasites (pathogenic and non-pathogenic) were identified. (4) Conclusions: Important pathogenic intestinal parasites were identified in fruits/vegetables commercially purchased in Maputo City. This fact must be taken into consideration when planning the management of these horticultural markets, in order to reduce the risk of contamination of fresh produce by intestinal parasites, and to prevent foodborne diseases.

Handling of Fresh Vegetables: Knowledge, Hygienic Behavior of Vendors, Public Health in Maputo Markets, Mozambique.

In developing countries, markets are the main supply of horticultural products to populations, but this can pose a public health challenge due to the risk of the fecal-oral transmission of gut pathogens. This transmission is strongly associated with inadequate public sanitation or low standards of personal and domestic hygiene, and their prevalence can cause gastrointestinal diseases, which are the third leading cause of death in Mozambique. This study aims at assessing the risk for public health of horticultural products supply chain, from the farmers-vendors to the consumers, in municipal markets in Maputo-City, Mozambique. Surveys (75) were conducted on vendors and an observational analysis was performed in the markets under study. The results showed that 62% of the vendors had access to water from boreholes or artisanal sources and the issue "access to water" was significantly different between markets (p = 0.004). Of the vendors who wash their products (53.3%), only 7.5% use tap-water for this purpose, with the difference in attitudes being statistically significant between vendors in the markets (p = 0.035). The majority (60.4%) said that vegetables and fruits can cause diseases due to pesticides and only 31.3% believe that the diseases may be related to poor hygiene. Despite the vendors' low knowledge of Good Hygiene Practices (GHP), we noticed that women have better practical assimilation of GHP when compared to men (p = 0.008). Although Maputo's markets are struggling to achieve quality hygiene standards in a reliable and sustainable manner, their resources are limited and significantly different (p = 0.044) from market to market, and this problem remains a concern for the public-health authorities of the city. In conclusion, the provision of adequate drinking water and sewage disposal systems, together with education for health of vendors, can reduce the risk of contamination of fresh food by the more common organisms causing diarrhea in children, including intestinal parasites.

Correction to: Plea for a standardized imaging approach to disorders of sex development in neonates: Consensus proposal from European Society of Paediatric Radiology task force.

The above article was published online with an incorrect author name.

Plea for a standardized imaging approach to disorders of sex development in neonates: consensus proposal from European Society of Paediatric Radiology task force.

This consensus article elaborated by the European Society for Paediatric Radiology task force on gastrointestinal and genitourinary imaging is intended to standardize the imaging approach in newborns with disorders of sex development. These newborns represent a difficult and stressful situation necessitating a multidisciplinary team approach. Imaging plays an important role in the work-up but needs to be optimized and customized to the patient. Ultrasound plays the central role in assessing the genital anatomy. The examination must be conducted in a detailed and systematic way. It must include transabdominal and transperineal approaches with adapted high-resolution transducers. The pelvic cavity, the genital folds, the inguinal areas and the adrenals must be evaluated as well as the rest of the abdominal cavity. A reporting template is proposed. The indications of magnetic resonance imaging and cysto- and genitography are discussed as well as they may provide additional information. Imaging findings must be reported cautiously using neutral wording as much as possible.

Prevalence and genotyping of Pneumocystis jirovecii in renal transplant recipients-preliminary report.

Pneumocystis jirovecii is an opportunistic fungus occurring in human lungs. The group at highest risk consists of HIV-infected and non-HIV-infected immunosuppressed individuals. In these patients, P. jirovecii infection may lead to Pneumocystis pneumonia; it may, however, persist also in an asymptomatic form. This study aimed to determine the prevalence of P. jirovecii and potential risk factors for infection in a group of renal transplant recipients and to characterize the genetic diversity of this fungus in the studied population. Sputum specimens from 72 patients were tested for presence of P. jirovecii using immunofluorescence microscopy, as well as nested PCR targeting the mtLSU rRNA gene. Genotyping involving analysis of four loci-mtLSU rRNA, CYB, DHPS, and SOD-was used to characterize the diversity of the detected organisms. Pneumocystis DNA was detected in eight (11.11%) patients. It has been shown that low eosinophil count and dual immunosuppressive treatment combining prednisone and calcineurin inhibitors are potential risk factors for colonization. Analysis of genotype distribution showed an association of the wild-type genotype of mtLSU rRNA with lower average age of patients and shorter time after kidney transplantation. Furthermore, CYB 2 genotype was detected only in patients with the ongoing prophylaxis regimen. In conclusion, renal transplant recipients are at risk of Pneumocystis colonization even a long time after transplantation. The present preliminary study identifies specific polymorphisms that appear to be correlated with certain patient characteristics and highlights the need for deeper investigation of these associations in renal transplant recipients.

Genotyping of Pneumocystis jirovecii in colonized patients with various pulmonary diseases.

Pneumocystis jirovecii is an opportunistic fungus causing Pneumocystis pneumonia primarily in immunosuppressed patients. However, immunocompetent individuals may become colonized and, as asymptomatic carriers, serve as reservoirs of the pathogen. Moreover, these asymptomatic carriers are at higher risk of developing pneumonia if favorable conditions occur. This study aimed to determine the prevalence of P. jirovecii in patients with various pulmonary diseases and to characterize the genetic diversity of organisms circulating in the studied population. Bronchial washing specimens from 105 patients were tested for presence of P. jirovecii using nested polymerase chain reaction (PCR) targeting the mtLSU rRNA gene, as well as immunofluorescence microscopy. Multilocus sequence typing involving analysis of three loci-mtLSU rRNA, CYB, and SOD-was used for genotyping analysis. P. jirovecii DNA was detected in 17 (16.2%) patients. Amplification of the SOD locus was successful only in five cases (29.4% of the positive patients), while mtLSU rRNA and CYB were genotyped in all positive samples. Therefore, combined genotypes were identified based only on mtLSU rRNA and CYB loci. Eight different genotypes were identified, with Pj 1 and Pj 2 being the most prevalent (29.4% of patients each). There was no statistical correlation between these genotypes and demographic or clinical data; however, we found that infection with mutant CYB strains occurred only in patients diagnosed with lung cancer. Of the potential predictors examined, only immunosuppressive treatment was significantly associated with colonization. In conclusion, patients with various respiratory diseases, especially when immunosuppressed, are at risk of Pneumocystis colonization.

Standardization of pediatric uroradiological terms: a multidisciplinary European glossary.

To promote the standardization of nephro-uroradiological terms used in children, the European Society of Paediatric Radiology uroradiology taskforce wrote a detailed glossary. This work has been subsequently submitted to European experts in pediatric urology and nephrology for discussion and acceptance to improve the quality of radiological reports and communication between different clinicians involved in pediatric urology and nephrology.

Standardization of pediatric uroradiological terms: A multidisciplinary European glossary.

To promote the standardization of nephro-uroradiological terms used in children, the European Society of Pediatric Radiology uroradiology taskforce wrote a detailed glossary. This work has been subsequently submitted to European experts in pediatric urology and nephrology for discussion and acceptance to improve the quality of radiological reports and communication among different clinicians involved in pediatric urology and nephrology.

European Society of Paediatric Radiology abdominal imaging task force recommendations in paediatric uroradiology, part IX: Imaging in anorectal and cloacal malformation, imaging in childhood ovarian torsion, and efforts in standardising paediatric uroradiology terminology.

At the occasion of the European Society of Paediatric Radiology (ESPR) annual meeting 2015 in Graz, Austria, the newly termed ESPR abdominal (gastrointestinal and genitourinary) imaging task force set out to complete the suggestions for paediatric urogenital imaging and procedural recommendations. Some of the last missing topics were addressed and proposals on imaging of children with anorectal and cloacal malformations and suspected ovarian torsion were issued after intense discussions and a consensus finding process that considered all evidence. Additionally, the terminology was adapted to fit new developments introducing the term pelvicalyceal dilatation/distension (PCD) instead of the sometimes misunderstood hydronephrosis. The present state of paediatric urogenital radiology was discussed in a dedicated minisymposium, including an attempt to adapt terminology to create a standardised glossary.

ESPR Uroradiology Taskforce--imaging recommendations in paediatric uroradiology, part VIII: retrograde urethrography, imaging disorder of sexual development and imaging childhood testicular torsion.

Three new consensus-based recommendations of the European Society of Paediatric Radiology Uroradiology Taskforce and the European Society of Urogenital Radiology Paediatric Working Group on paediatric uroradiology are presented. One deals with indications and technique for retrograde urethrography, one with imaging in the work-up for disorders of sexual development and one with imaging workflow in suspected testicular torsion. The latter is subdivided to suggest a distinct algorithm to deal with testicular torsion in neonates. These proposals aim to outline effective imaging algorithms to optimise diagnostic accuracy and to harmonize diagnostic imaging among institutions and practitioners.

Molecular characterisation of Cryptosporidium (Apicomplexa) in children and cattle in Romania.

To investigate the transmission of species of Cryptosporidium Tyzzer, 1907 in Timis County, Romania, 48 isolates of Cryptosporidium coccidia from 11 children, 29 calves and eight pigs were characterised by molecular analysis of two loci (SSU rRNA and 60-kDa glycoprotein gene). Overall, 22 isolates were amplified and sequence analyses revealed that all isolates were Cryptosporidium parvum Tyzzer, 1912. Two subtype families were identified, IIa and IId. Subtype IIdA22G1 (n = 4) was the single C. parvum subtype found in children. Subtypes found in calves included IIdA27G1 (n = 8), a novel subtype, IIdA25G1 (n = 5), IIdA22G1 (n = 2), IIdA21G1a (n = 1), and IIaA16G1R1 (n = 1). Subtype IIdA26G1 was found in a pig. These results were significantly different from previous Romanian reports, as the five subtypes of family IId identified in this study were never identified previously in this country. Thus, cattle may be a source of Cryptosporidium infections for humans and the transmission dynamics of C. parvum in Romania is more complex than previously believed.

ESPR uroradiology task force imaging recommendations in paediatric uroradiology, part VII: standardised terminology, impact of existing recommendations, and update on contrast-enhanced ultrasound of the paediatric urogenital tract.

Our purpose is to harmonise and standardise terminology in paediatric uroradiology, to provide and update recommendations for contrast-enhanced US to standardise imaging and encourage further research, and to assess the impact of the existing recommendations in paediatric urogenital imaging. Based on thorough review of literature and variable practice at several centres and after discussion within urogenital imaging groups as well as with other subspecialties, we propose a standardisation of terminology in urogenital imaging. An update with recommendations on paediatric contrast-enhanced US has been issued based on available literature and reports. Finally, a questionnaire has been used to assess the knowledge, applicability and usefulness of, and the adherence to existing recommendations of the European Society of Paediatric Radiology (ESPR) Uroradiology Task Force. In conclusion, the ESPR is working to improve patient safety and optimise paediatric urogenital imaging. Standardisation of terminology and provision of updated knowledge on contrast-enhanced US in childhood will contribute to this task, ideally reducing the need for invasive or radiating imaging. Not all existing recommendations are commonly known, which limits adherence to these recommendations and the availability of comparable data and evidence for future adaptation of imaging strategies in paediatric uroradiology.

Cryptosporidium spp., Giardia duodenalis, Enterocytozoon bieneusi and other intestinal parasites in young children in Lobata province, Democratic Republic of São Tomé and Principe.

Rare systemic studies concerning prevalence of intestinal parasites in children have been conducted in the second smallest country in Africa, the Democratic Republic of São Tomé and Príncipe. Fecal specimens from 348 children (214 in-hospital attending the Aires de Menezes Hospital and 134 from Agostinho Neto village) in São Tome Island were studied by parasitological and molecular methods. Of the 134 children from Agostinho Neto, 52.2% presented intestinal parasites. 32.1% and 20.2% of these children had monoparasitism and polyparasitism, respectively. Ascaris lumbricoides (27.6%), G. duodenalis (7.5%), T. trichiura (4.5%) and Entamoeba coli (10.5%) were the more frequent species identified in the children of this village. Giardia duodenalis (7.5%) and E. bieneusi (5.2%) were identified by PCR. Nested-PCR targeting G. duodenalis TPI identified Assemblage A (60%) and Assemblage B (40%). The E. bieneusi ITS-based sequence identified genotypes K (57.1%), KIN1 (28.6%) and KIN3 (14.3%). Among the 214 in-hospital children, 29.4% presented intestinal parasites. In 22.4% and 7.0% of the parasitized children, respectively, one or more species were concurrently detected. By microscopy, A. lumbricoides (10.3%) and Trichiuris trichiura (6.5%) were the most prevalent species among these children, and Cryptosporidium was detected by PCR in 8.9% of children. GP60 locus analysis identified 6.5% of C. hominis (subtypes IaA27R3 [35.7%], IaA23R3 [14.3%], IeA11G3T3 [28.6%] and IeA11G3T3R1 [21.4%]) and 2.3% of C. parvum (subtypes IIaA16G2R1 [20.0%], IIaA15G2R1 [20.0%], IIdA26G1 [40.0%] and IIdA21G1a [20.0%]). G. duodenalis and E. bieneusi were identified in 0.5% and 8.9% of the in-hospital children, respectively. G. duodenalis Assemblage B was characterized. The E. bieneusi genotypes K (52.6%), D (26.4%), A (10.5%) and KIN1 (10.5%) were identified. Although further studies are required to clarify the epidemiology of these infectious diseases in this endemic region the significance of the present results highlights that it is crucial to strength surveillance on intestinal pathogens.

Gastrointestinal ultrasound in neonates, infants and children.

Today US plays an important and increasing role in the assessment of many, partially age-specific conditions in the GI tract in neonates, infants and children. Knowledge of the potential capabilities of US and its restrictions together with a skillful performance of GI US examination can provide essential anatomic and functional diagnostic information in many pediatric GI disorders. The aim of this review is to highlight the potential of ultrasound (US) in the evaluation of the gastrointestinal (GI) tract in neonates, infants and children. Basic and potential applications of modern US tools in pediatric GI tract are addressed, the GI US examination technique is discussed - including some common and/or typical clinical applications of and indications for US.

ESPR uroradiology task force and ESUR Paediatric Work Group--Imaging recommendations in paediatric uroradiology, part VI: childhood renal biopsy and imaging of neonatal and infant genital tract. Minutes from the task force session at the annual ESPR Meeting 2012 in Athens on childhood renal biopsy and imaging neonatal genitalia.

The European Society of Paediatric Radiology Uroradiology Task Force and the ESUR Paediatric Work Group jointly publish guidelines for paediatric urogenital imaging. Two yet unaddressed topics involving patient safety and imaging load are addressed in this paper: renal biopsy in childhood and imaging of the neonatal genital tract, particularly in girls. Based on our thorough review of literature and variable practice in multiple centers, procedural recommendations are proposed on how to perform renal biopsy in children and how to approach the genital tract in (female) neonates. These are statements by consensus due to lack of sufficient evidence-based data. The procedural recommendation on renal biopsy in childhood aims at improving patient safety and reducing the number of unsuccessful passes and/or biopsy-related complications. The recommendation for an imaging algorithm in the assessment of the neonatal genital tract focuses on the potential of ultrasonography to reduce the need for more invasive or radiating imaging, however, with additional fluoroscopy or MRI to be used in selected cases. Adherence to these recommendations will allow comparable data and evidence to be generated for future adaptation of imaging strategies in paediatric uroradiology.

Therapeutic potential of caspofungin combined with trimethoprim-sulfamethoxazole for pneumocystis pneumonia: a pilot study in mice.

Pneumocystis pneumonia (PcP) is a major cause of mortality and morbidity in immunocompromised patients. There are limited alternative therapeutic choices to trimethoprim-sulfamethoxazole (TMP-SMX) which is the standard first line therapy/prophylaxis for PcP. The efficacy of low doses of caspofungin and caspofungin in association with TMP-SMX standard-prophylactic dose was evaluated in an experimental model of Pneumocystis. Susceptibility of Pneumocystis spp. to low doses of caspofungin and caspofungin/TMP-SMX was evaluated in Balb/c immunosuppressed mice, infected intranasally with P. murina. Caspofungin was administered once daily at 0.1 mg/kg, 0.05 mg/kg, and 0.001 mg/kg and TMP-SMX was administered by oral gavage (12.25 mg/62.5 mg/day), for 21 days. Efficacy was calculated based on the reduction in organism burden determined through quantitative fluorescent-based real-time PCR (qPCR). Serum ß-1,3-D-glucan was measured as an additional marker of infection. The present data showed that caspofungin demonstrated anti-Pneumomocystis effect. However, the doses administrated were too low to achieve Pneumocystis eradication, which suggests that echinocandin treatment should not be administrated as mono-therapy. After 21 days of treatment, P. murina was not detected in the lungs of mice with either TMP-SMX or caspofungin/TMP-SMX. The results showed that, even at the lowest concentrations tested, the efficacy of caspofungin in association with TMP-SMX was higher than the efficacy of either drug used alone. The administration of caspofungin/TMP-SMX was at least 1.4 times more effective against P. murina infection than TMP-SMX used alone. The most promising result was achieved with the combination of caspofungin 0.05 mg/kg/day with TMP-SMX 12.5 mg-62.5 mg/day, which reduced the parasite burden to undetectable levels immediately at the 14(th) day of treatment, showing a highly marked anti-Pneumomocystis effect. These data suggest that the administration of low doses of caspofungin in combination with low doses of TMP-SMX may provide an improved treatment protocol for Pneumocystis infection clearance.

Epidemiology of Enterocytozoon bieneusi Infection in Humans.

A review was conducted to examine published works that focus on the complex epidemiology of Enterocytozoon bieneusi infection in humans. Studies on the prevalence of these emerging microsporidian pathogens in humans, in developed and developing countries, the different clinical spectra of E. bieneusi intestinal infection in children, in different settings, and the risk factors associated with E. bieneusi infection have been reviewed. This paper also analyses the impact of the recent application of PCR-based molecular methods for species-specific identification and genotype differentiation has had in increasing the knowledge of the molecular epidemiology of E. bieneusi in humans. The advances in the epidemiology of E. bieneusi, in the last two decades, emphasize the importance of epidemiological control and prevention of E. bieneusi infections, from both the veterinary and human medical perspectives.