Growth and gut comfort of healthy term infants exclusively fed with a partially hydrolysed protein-based infant formula: a randomized controlled double-blind trial.

Objective: This study aimed to investigate growth and gut comfort of healthy infants fed with a partially hydrolysed cow's milk protein-based infant formula (pHF) compared to a standard intact cow's milk protein-based formula (IPF). Methods: A double-blind, multi-center, randomized, controlled trial was performed. Healthy full-term, exclusively formula-fed infants (n = 345), aged ≤28 days were allocated to consume either a pHF (n = 173) or an IPF (n = 172) until the age of 17 weeks. The primary outcome was equivalence of weight gain (g/d) until the age of 17 weeks. The secondary outcomes were equivalence of other growth parameters, i.e., infants' weight, length, head circumference, body mass index (BMI) and anthropometric Z-scores, while tertiary outcomes were gut comfort, formula intake, and adverse events (AEs). Results: Overall, 288 infants completed the study (pHF group: 138, IPF group: 150). No differences were observed between the two groups in weight gain (g/d) during the three-months intervention [p = 0.915 for the Per Protocol (PP) population]. The 90% CI was [-1.252 to 1.100] being within the pre-defined equivalence margin of ±3.0 g/d. Similar findings were observed in the Full Analysis Set (FAS) and the sensitivity analysis. Regarding the secondary outcomes, no differences over the intervention period were shown between the two groups in both the PP and FAS analysis sets. Average Z-scores were in the normal range based on World Health Organization (WHO) growth standards for both groups at all time points in both analysis sets. Stool consistency, amount, and colour were different in the two groups. No differences were observed in gut comfort, stool frequency, and formula intake, between the two groups. In total 14 AEs and 22 serious adverse events (SAEs) were reported of which 15 (12%) and 1 (5%) were considered as (possibly) related to the study product, respectively. Conclusions: The study demonstrates that the consumption of pHF results in adequate infant growth, equivalent to that of infants consuming IPF. Furthermore, the overall gut comfort was comparable between the two groups. Therefore, it can be concluded that the pHF is safe for and well tolerated by healthy infants. Clinical Trial Registration: https://clinicaltrials.gov/study/NCT05757323?id=NCT05757323&rank=1, identifier (NCT05757323).

A cross-sectional study on stool- and gastrointestinal-related outcomes of Mexican infants consuming different formulae.

BACKGROUND: Immaturities present at birth, such as in the gut microbiome and digestive, nervous, and immune system, resolve with time. Nevertheless, this may result in mild digestive symptoms early in life, particularly in formula-fed infants. Formula composition and processing may impact this discomfort. This study therefore aimed to assess stool characteristics and gastrointestinal symptoms of healthy infants fed different formulae. METHODS: A multicenter, cross-sectional, observational trial was performed in Mexico between November 2019 and January 2022, where exclusively formula-fed infants (n = 342, aged 1-4 months) were studied in four groups based on their existing formula use. Feeding was continued per practice following label instructions. For 7 days, parents/caregivers were requested to record fecal characteristics, using the Amsterdam Infant Stool Scale, and rate gastrointestinal symptoms. Stool samples were collected to determine pH, dry matter content, and fecal calprotectin levels. RESULTS: Most infants had a soft/formed stool consistency, although odds for hard stools were different between groups. Gastrointestinal symptom scores revealed significant differences for burping and diarrhea, while other symptoms did not differ between groups. No significant differences between groups were found for stool frequency, dry matter content, and fecal pH. Although calprotectin was within the expected healthy ranges, significant differences among groups were seen. Furthermore, calprotectin significantly correlated with the severity of the gastrointestinal symptoms burping, flatulence, abdominal distension, and diarrhea. CONCLUSIONS: Differences in stool characteristics and specific differences in gastrointestinal symptoms were observed between different formula brand users. This may potentially be explained by the different composition and processing of the formulae, although there are multiple factors that influence the assessed outcomes. TRIAL REGISTRATION: The study was registered in the Netherlands Trial Registry (NL7805), linked to https://trialsearch.who.int/ , on 11/06/2019.

Symptom's resolution and growth outcome of children with cow's milk protein allergy consuming two hydrolyzed formulas: A retrospective study in Mexico.

Background: Cow's milk protein allergy (CMPA) is the leading cause of food allergy in infants and young children. An extensively hydrolyzed formula (eHF) is the first choice of dietary management, however, not all of them have similar peptide profiles and degree of hydrolysis. The aim of this retrospective study was to investigate the use of two commercially available infant formulas in the clinical management of CMPA in Mexico in terms of symptoms' resolution and growth trajectories. Methods: Medical records of 79 subjects from four sites in Mexico were included to retrospectively evaluate the trajectory of atopic dermatitis, other symptoms of cow's milk protein allergy and growth outcomes. The study formulas were based on hydrolyzed whey protein (eHF-W) and hydrolyzed casein protein (eHF-C). Results: Medical records of 79 patients were enrolled, 3 were excluded from analysis based on previous formula consumption. Seventy-six children with confirmed CMPA based on skin prick test and/or serum specific IgE levels were included in the analysis. 82% of patients (n = 65) consumed the eHF-C, reflecting the doctors' preference for formulas with a higher grade of hydrolysis and the high incidence of positive reactions to beta-lactoglobulin amongst subjects. During their first visit to the doctors, 55% of the subjects consuming the casein-based formula and 45% of subjects consuming the whey-based formula presented with mild or moderate dermatological symptoms. Other frequently reported symptoms included respiratory issues, enteropathies and colitis which improved during the consumption of both formulas. All CMPA-related symptoms showed improvement during formula consumption. During the period of retrospective observation, growth significantly improved for both groups. Conclusion: Consumption of eHF-C and eHF-W effectively improved symptoms' resolution and growth outcomes among children with CMPA in Mexico. More preference was reported towards eHF-C due to its hydrolysate profile and lack of b-lactoglobulin. Trial registration: The study was registered at ClinicalTrials.gov: NCT04596059.

The use of two Comfort Young Child Formulas in the dietary management of toddlers with functional constipation: a randomized controlled trial.

BACKGROUND: Pharmacological intervention with laxatives is the conventional treatment for functional constipation (FC). Data to support the dietary management of FC is lacking. This study compared the efficacy of two Comfort young child formulas (YCFs) with regards to the maintenance of healthy stooling parameters in toddlers with a history of constipation. It was registered in the Netherlands Trial Registry [identifier: NL7420 (NTR7653)], registration date 20/09/2018. METHODS: Ninety-five healthy toddlers, aged 12 to 32 months, diagnosed with FC (Rome III criteria) were randomized to receive one of two study formulas after pharmacological treatment. For the first month of the intervention, subjects received a laxative in a decreasing maintenance dose alongside a test or control formula (maintenance phase). Subsequently, subjects only consumed formula for another month (post-maintenance phase). Stooling parameters were obtained weekly using the Bristol Stool Scale and the modified Rome III Questionnaire on Paediatric Gastrointestinal Symptoms for infants and toddlers. Differences in percentages of hard stools (primary outcome) and other stooling parameters were analysed using analysis of covariance and Chi-Square methods. RESULTS: Both formulas resulted in similar overall percentage of hard stools during the intervention period, respectively 5.02% in the test and 2.99% in the control group (n.s.). In the test group, percentages dropped from 7.11% at the end of the maintenance phase, to 3.92% at the end of the post-maintenance phase. In contrast, the percentage of hard stools in the control group was similar at the end of the maintenance (3.18%) and post-maintenance phase (2.83%; n.s.). No difference was found in the overall stool frequency between groups. At the end of the maintenance phase, only 22% and 19% of toddlers consuming the test and control formulae, respectively, met 2 or more of the criteria for FC. At the end of the study, this percentage of subjects decreased further to 9% in the test group, which tended to be lower compared to the 21% found in the control (p = 0.107). No laxative use was reported in either study group during the post-maintenance phase. CONCLUSION: Both Comfort YCF support the maintenance of improved stooling over time in toddlers with a history of constipation. The percentage of subjects suffering from functional constipation tended to be lower after the intervention period when receiving the formula with intact protein.

Effect of a Fortified Dairy-Based Drink on Micronutrient Status, Growth, and Cognitive Development of Nigerian Toddlers- A Dose-Response Study.

Malnutrition results in a high prevalence of stunting, underweight, and micronutrient deficiencies. This study investigated the effect of a multi-nutrient fortified dairy-based drink on micronutrient status, growth, and cognitive development in malnourished [height-for-age z-score (HAZ) and/or weight-for-age z-score (WAZ) < -1 SD and >-3 SD] Nigerian toddlers (n = 184, 1-3 years). The product was provided in different daily amounts (200, 400, or 600 ml) for 6 months. At baseline and endline, venous blood and urine samples were collected to determine micronutrient status. Bodyweight, height, waist, and head circumference were measured, and corresponding Z-scores were calculated. The Bayley-III Screening Test was used to classify the cognitive development of the children. In a modified per-protocol (PP) population, the highest prevalence's of micronutrient deficiencies were found for vitamin A (35.5%) and selenium (17.9%). At endline, there were no significant improvements in iodine, zinc, vitamin B12, and folate status in any of the three groups. Regarding vitamin D status (25OHD), consumption of 600 and 400 ml resulted in an improved status as compared to baseline, and in a difference between the 600- and 200-ml groups. Consumption of 600 ml also increased vitamin A and selenium status as compared to baseline, but no differences were found between groups. Within the groups, WAZ, weight-for-height z-score (WHZ), and BMI-for-age z-score (BAZ) improved, but without differences between the groups. For HAZ, only the 600 ml group showed improvement within the group, but it was not different between groups. For the absolute weight, height, and head circumference only trends for differences between groups were indicated. Cognition results did not differ between the groups. Within groups, all showed a decline in the per cent of competent children for receptive language. To study the effects of a nutritional intervention on linear growth and cognition, a longer study duration might be necessary. Regarding the improvement of micronutrient status, 600 ml of fortified dairy-based drink seems most effective. Clinical Trial Registration: https://clinicaltrials.gov/ct2/show/NCT03411590?term=NCT03411590.&draw=2&rank=1, identifier: NCT03411590.

Prebiotic Galacto-Oligosaccharides Impact Stool Frequency and Fecal Microbiota in Self-Reported Constipated Adults: A Randomized Clinical Trial.

Constipation is a major issue for 10-20% of the global population. In a double-blind randomized placebo-controlled clinical trial, we aimed to determine a dose-response effect of galacto-oligosaccharides (GOS) on stool characteristics and fecal microbiota in 132 adults with self-reported constipation according to Rome IV criteria (including less than three bowel movements per week). Subjects (94% females, aged: 18-59 years) received either 11 g or 5.5 g of BiotisTM GOS, or a control product, once daily for three weeks. Validated questionnaires were conducted weekly to study primarily stool frequency and secondary stool consistency. At base- and endline, stool samples were taken to study fecal microbiota. A trend towards an increased stool frequency was observed after the intervention with 11 g of GOS compared to control. While during screening everybody was considered constipated, not all subjects (n = 78) had less than three bowel movements per week at baseline. In total, 11 g of GOS increased stool frequency compared to control in subjects with a low stool frequency at baseline (≤3 bowel movements per week) and in self-reported constipated adults 35 years of age or older. A clear dose-response of GOS was seen on fecal Bifidobacterium, and 11 g of GOS significantly increased Anaerostipes hadrus. In conclusion, GOS seems to be a solution to benefit adults with a low stool frequency and middle-aged adults with self-reported constipation.

Multi-Nutrient Fortified Dairy-Based Drink Reduces Anaemia without Observed Adverse Effects on Gut Microbiota in Anaemic Malnourished Nigerian Toddlers: A Randomised Dose-Response Study.

Prevalence of anaemia among Nigerian toddlers is reported to be high, and may cause significant morbidity, affects brain development and function, and results in weakness and fatigue. Although, iron fortification can reduce anaemia, yet the effect on gut microbiota is unclear. This open-label randomised study in anaemic malnourished Nigerian toddlers aimed to decrease anaemia without affecting pathogenic gut bacteria using a multi-nutrient fortified dairy-based drink. The test product was provided daily in different amounts (200, 400 or 600 mL, supplying 2.24, 4.48 and 6.72 mg of elemental iron, respectively) for 6 months. Haemoglobin, ferritin, and C-reactive protein concentrations were measured to determine anaemia, iron deficiency (ID) and iron deficiency anaemia (IDA) prevalence. Faecal samples were collected to analyse gut microbiota composition. All three dosages reduced anaemia prevalence, to 47%, 27% and 18%, respectively. ID and IDA prevalence was low and did not significantly decrease over time. Regarding gut microbiota, Enterobacteriaceae decreased over time without differences between groups, whereas Bifidobacteriaceae and pathogenic E. coli were not affected. In conclusion, the multi-nutrient fortified dairy-based drink reduced anaemia in a dose-dependent way, without stimulating intestinal potential pathogenic bacteria, and thus appears to be safe and effective in treating anaemia in Nigerian toddlers.

The Effect of Partly Replacing Vegetable Fat with Bovine Milk Fat in Infant Formula on Postprandial Lipid and Energy Metabolism: A Proof-of-principle Study in Healthy Young Male Adults.

SCOPE: Infant formula (IF) uses besides vegetable fats also bovine milk fat, which differs in triacylglycerol (TAG) structure. Furthermore, it differs in fatty acid (FA) composition. Whether changing fat source in IF affects postprandial energy metabolism, lipemic response, and blood lipid profile is unknown. METHODS AND RESULTS: A proof-of-principle study, with a randomized controlled double-blind cross-over design, is conducted. Twenty healthy male adults consumed drinks with either 100% vegetable fat (VEG) or 67% bovine milk fat and 33% vegetable fat (BOV), on 2 separate days. For a detailed insight in the postprandial responses, indirect calorimetry is performed continuously, and venous blood samples are taken every 30 min, until 5 h postprandially. No differences in postprandial energy metabolism, serum lipids, lipoprotein, or chylomicron concentrations are observed between drinks. After consumption of VEG-drink, C18:2n-6 in serum increased. Observed differences in chylomicron FA profile reflect differences in initial FA profile of test drinks. Serum ketone bodies concentrations increase following consumption of BOV-drink. CONCLUSIONS: The use of bovine milk fat in IF does neither affect postprandial energy metabolism nor lipemic response in healthy adults, but alters postprandial FA profiles and ketone metabolism. Whether the exact same effects occur in infants requires experimental verification.

Application of Volatile Organic Compound Analysis in a Nutritional Intervention Study: Differential Responses during Five Hours Following Consumption of a High- and a Low-Fat Dairy Drink.

SCOPE: Exhaled volatile organic compounds (VOCs) are a possible relevant target for noninvasive assessment of metabolic responses. Using a breathomics approach, it is aimed to explore whether lipid intake influences VOC profiles in exhaled air, and to obtain insight in intra- and interindividual variations. METHODS AND RESULTS: Three human interventions are performed. In the first, 12 males consume a high-fat drink on three study days. In the second, 12 males receive a high- and a low-fat drink on 6 days. In the third, three volunteers consume the high-fat drink again for tentative compound identification. Participants are asked to exhale, for 5 h postprandial with 15-20 min intervals, into a proton-transfer-reaction mass spectrometer, and VOCs in exhaled air are measured. Consumption of a drink alters the VOC profile, with considerable interindividual variation and quantitative intraindividual differences between days. Consumption of two different drinks results in a distinct VOC profile, caused by several specific m/z values. Most of these compounds are identified as being related to ketone body formation and lipid oxidation, showing an increase in high- versus low-fat drink. CONCLUSION: Exhaled VOCs have the potential to assess differences in metabolic responses induced by nutrition, especially when day-to-day variation can be minimized.

Correction: Free fatty acid release from vegetable and bovine milk fat-based infant formulas and human milk during two-phase in vitro digestion.

Correction for 'Free fatty acid release from vegetable and bovine milk fat-based infant formulas and human milk during two-phase in vitro digestion' by Jeske H. J. Hageman et al., Food Funct., 2019, 10, 2102-2113.

Free fatty acid release from vegetable and bovine milk fat-based infant formulas and human milk during two-phase in vitro digestion.

BACKGROUND: Bovine milk fat is increasingly used in infant formula (IF). The triacylglycerol (TAG) structure of bovine milk fat might be beneficial for digestion and absorption. We investigated the release of fatty acids (FAs) of IF containing different fat blends and compared this to human milk. METHODS: Fresh human milk was sampled and two IFs were produced; one containing 100% vegetable fat (IF1) and one with 67% bovine milk fat and 33% vegetable fat (IF2). Using a static in vitro infant digestion model, consisting of a gastric and duodenal phase, the time dependent release of individual free fatty acids (FFA) was studied and analysed using GC-MS, and residual TAG levels were determined by GC-FID. RESULTS: Human milk and the IFs showed comparable total FA release. In the gastric phase, 4-11% of lipolysis occurred, and mainly short (SCFA)- and medium chain fatty acids (MCFA) were released. In the duodenal phase, lipolysis proceeded with release of C4:0 but was marked by a fast release of long-chain fatty acids (LCFA). The digestion of the IFs resulted in different FFA profiles during and at the end of digestion. IF2 gave more release of C4:0-C11:0, which reflects the FA composition of bovine milk. CONCLUSION: The addition of bovine milk fat to IF resulted in a total FA release comparable to an IF with only vegetable fat and human milk. However, it did lead to a different time-dependent release of individual FAs, which might result in differences in absorption and other health effects in vivo.

Virus-directed formation of electrocatalytically active nanoparticle-based Co3O4 tubes.

Spinel-type Co3O4 finds applications in a wide range of fields, including clean energy conversion, where nanostructured Co3O4 may provide a cost-efficient alternative to platinum- and iridium-based catalysts for electrocatalytic water-splitting. We here describe a novel strategy in which basic cobalt carbonate - a precursor to Co3O4 - is precipitated as sheet-like structures and microspheres covered with fine surface protrusions, via ammonium carbonate decomposition at room temperature. Importantly, these mild reaction conditions enable us to employ bio-inspired templating approaches to further control the mineral structure. Rod-like tobacco mosaic viruses (TMV) were used as biotemplates for mineral deposition, where we profit from the ability of Co(ii) ions to mediate the ordered assembly of the virus nanorods to create complex tubular superstructures of TMV/ basic cobalt carbonate. Calcination of these tubules is then achieved with retention of the gross morphology, and generates a hierarchically-structured solid comprising interconnected Co3O4 nanoparticles. Evaluation of these Co3O4 materials as electrocatalysts for the oxygen evolution reaction (OER) demonstrates that the activity of Co3O4 prepared by calcination of ammonia diffusion-grown precursors in both, the absence or presence of TMV exceeds that of a commercial nanopowder.

Somatic homologous recombination in plants is promoted by a geminivirus in a tissue-selective manner.

Four transgenic Arabidopsis thaliana lines carrying different reporter gene constructs based on split glucuronidase genes were used to monitor the frequency of somatic homologous recombination after geminivirus infections. Euphorbia mosaic virus and Cleome leaf crumple virus were chosen as examples, because they induce only mild symptoms and are expected to induce less general stress responses than other geminiviruses. After comparing the different plant lines and viruses as well as optimizing the infection procedure, Euphorbia mosaic virus enhanced recombination rates significantly in the transgenic reporter line 1445. The effect was tissue-specific in cells of the leaf veins as expected for this phloem-limited virus. The advantage for geminiviruses to activate a general recombination pathway is discussed with reference to an increased fitness by generating virus recombinants which have been observed frequently as an epidemiologic driving force.

The impact of dietary long-chain polyunsaturated fatty acids on respiratory illness in infants and children.

Increasing evidence suggests that intake of long-chain polyunsaturated fatty acids (LCPUFA), especially omega-3 LCPUFA, improves respiratory health early in life. This review summarizes publications from 2009 through July 2012 that evaluated effects of fish, fish oil or LCPUFA intake during pregnancy, lactation, and early postnatal years on allergic and infectious respiratory illnesses. Studies during pregnancy found inconsistent effects in offspring: two showed no effects and three showed protective effects of omega-3 LCPUFA on respiratory illnesses or atopic dermatitis. Two studies found that infants fed breast milk with higher omega-3 LCPUFA had reduced allergic manifestations. Earlier introduction of fish improved respiratory health or reduced allergy in four studies. Three randomized controlled trials showed that providing LCPUFA during infancy or childhood reduced allergy and/or respiratory illness while one found no effect. Potential explanations for the variability among studies and possible mechanisms of action for LCPUFA in allergy and respiratory disease are discussed.

Conformation-selective methylation of geminivirus DNA.

Geminiviruses with small circular single-stranded DNA genomes replicate in plant cell nuclei by using various double-stranded DNA (dsDNA) intermediates: distinct open circular and covalently closed circular as well as heterogeneous linear DNA. Their DNA may be methylated partially at cytosine residues, as detected previously by bisulfite sequencing and subsequent PCR. In order to determine the methylation patterns of the circular molecules, the DNAs of tomato yellow leaf curl Sardinia virus (TYLCSV) and Abutilon mosaic virus were investigated utilizing bisulfite treatment followed by rolling circle amplification. Shotgun sequencing of the products yielded a randomly distributed 50% rate of C maintenance after the bisulfite reaction for both viruses. However, controls with unmethylated single-stranded bacteriophage DNA resulted in the same level of C maintenance. Only one short DNA stretch within the C2/C3 promoter of TYLCSV showed hyperprotection of C, with the protection rate exceeding the threshold of the mean value plus 1 standard deviation. Similarly, the use of methylation-sensitive restriction enzymes suggested that geminiviruses escape silencing by methylation very efficiently, by either a rolling circle or recombination-dependent replication mode. In contrast, attempts to detect methylated bases positively by using methylcytosine-specific antibodies detected methylated DNA only in heterogeneous linear dsDNA, and methylation-dependent restriction enzymes revealed that the viral heterogeneous linear dsDNA was methylated preferentially.

Ambivalent effects of defective DNA in beet curly top virus-infected transgenic sugarbeet plants.

Beet curly top virus (BCTV) limits sugarbeet production considerably. Previous studies have shown that infections are associated with the generation of defective DNAs (D-DNA) which may attenuate symptoms. Transgenic sugarbeet lines were established carrying a partial direct repeat construct of D-DNA in order to examine whether they are useful as a means of generating tolerance against BCTV. Thirty four independent transgenic lines were challenged. Viral full-length and D-DNAs were monitored by polymerase chain reaction (PCR) or rolling circle amplification (RCA) and restriction fragment length polymorphism (RFLP). The differential accumulation of both DNA species was compared with symptom severity during the course of infection. RCA/RFLP allowed the discrimination of two D-DNA classes which were either derived from the transgenic construct (D(0)) or had been generated de novo (D(n)). The statistical analysis of the results showed that the presence of D(0)-DNA correlated with increased symptom severity, whereas D(n)-DNAs correlated with attenuated symptoms.

The first DNA 1-like alpha satellites in association with New World begomoviruses in natural infections.

From Brazilian weeds with typical symptoms of a geminivirus infection, the DNAs of two new virus species, two new strains with two variants of already known bipartite begomoviruses were sequenced. Moreover, the first two DNA 1-like satellites (alpha satellites) occurring naturally in the New World were identified. They are related to nanoviral DNA components and show a typical genome organization with one open reading frame coding potentially for a replication-associated protein (Rep), a conserved hairpin structure, and an A-rich region. After coinoculation with their helper begomoviruses (Euphorbia mosaic virus, EuMV or Cleome leaf crumple virus, ClLCrV) the satellite DNAs were transmitted to experimental and natural host plants. Three of the begomovirus isolates (EuMV and ClLCrV) infected Arabidopsis thaliana plants, induced mild symptoms, and one of these (ClLCrV) transreplicated the satellite efficiently. As a result, several novel tools for molecular analyses of this important model plant are provided.

The complete nucleotide sequence of a new bipartite begomovirus from Brazil infecting Abutilon.

The complete nucleotide sequence of Abutilon mosaic Brazil virus (AbMBV), a new bipartite begomovirus from Bahia, Brazil, is described and analyzed phylogenetically. Its DNA A is most closely related to those of Sida-infecting begomoviruses from Brazil and forms a phylogenetic cluster with pepper- and Euphorbia-infecting begomoviruses from Central America. The DNA B component forms a cluster with different Sida- and okra-infecting begomoviruses from Brazil. Both components are distinct from those of the classical Abutilon mosaic virus originating from the West Indies. AbMBV is transmissible to Nicotiana benthamiana and Malva parviflora by biolistics of rolling-circle amplification products and induces characteristic mosaic and vein-clearing symptoms in M. parviflora.

Genomic diversity of sweet potato geminiviruses in a Brazilian germplasm bank.

Begomoviruses cause major diseases of sweet potato worldwide impairing considerably the yields of this important food staple. Since sweet potato plants are vegetatively propagated and globally transported, they are prone to accumulate and disseminate geminiviruses. Effective diagnostic tools are, therefore, desirable. We studied the genomic diversity of geminiviruses present in naturally-infected sweet potato accessions belonging to a Brazilian germplasm bank collection. Fifty-five samples from different sweet potato accessions displaying geminivirus-like symptoms were analyzed by combining rolling circle amplification (RCA) with restriction fragment length polymorphism (RFLP) and sequencing. The restriction enzyme MspI (HpaII) revealed diverse band patterns in 55 samples and digestion with BamHI, SstI or PstI resulted in full-length sweet potato geminivirus DNAs of about 3 kb in 46 samples. In addition, smaller fragments were identified as either viral "Defective DNAs" (D-DNAs) or mitochondrial plasmid DNAs. The diversity of sweet potato-associated geminiviruses was found to be very high under Brazilian conditions. Representative viral full-length DNAs have been cloned and sequenced yielding two new tentative species, three strains and several variants of previously described sweet potato geminiviruses. Sequence comparisons identified footprints of recombination in their genomes underscoring the risk of generating new geminiviruses in vegetatively propagated germplasm bank material. The sites of recombination were found in conjunction with predicted hairpin structures. We propose diagnostic routines to screen germplasm bank material for geminiviruses by the rapid and reliable RCA/RFLP as the technique of choice.

Geminiviruses.

Plant pathogenic geminiviruses have been proliferating worldwide and have, therefore, attracted considerable scientific interest during the past three decades. Current knowledge concerning their virion and genome structure, their molecular biology of replication, recombination, transcription, and silencing, as well as their transport through plants and dynamic competition with host responses are summarized. The topics are chosen to provide a comprehensive introduction for animal virologists, emphasizing similarities and differences to the closest functional relatives, polyomaviruses and circoviruses.