Long-term sequelae of sporadic cryptosporidiosis: a follow-up study.

To determine the frequency of occurrence of sequelae following cryptosporidiosis. A follow-up study was performed during a case-control study for sporadic cryptosporidiosis in the Netherlands (2013-2016). Cryptosporidiosis cases were invited to complete a follow-up questionnaire 4 months after diagnosis. Using a case-crossover study design, we compared the frequencies of reported symptoms 4 months after the acute phase to those reported 4 months before the onset of illness and during illness. Frequencies of symptoms in the pre- to post-infection phases were also compared with those of a population control group. Cryptosporidium species-specific effects were also studied. Logistic regression was used to calculate adjusted odds ratios (aOR) for symptoms occurrence. Of the 731 available cases, 443 (60%) responded and 308 (42%) could be included in the follow-up study. The median age was 26 years (range 1-80); 58% were female; 30% were infected with C. hominis and 70% with C. parvum. Compared to before illness, cases were significantly more likely to report dizziness (OR = 2.25), headache (OR = 2.15), fatigue (OR = 2.04), weight loss (OR = 1.82), diarrhoea (OR = 1.50), abdominal pain (OR = 1.38) or joint pain (OR = 1.84). However, symptoms of joint pain and headache occurred among cases after illness at a rate that was not significantly different from that observed in the general population. There were no significant differences in post-infection symptom occurrence between C. hominis and C. parvum. The disease burden of cryptosporidiosis extends beyond the acute phase of the infection, with cases reporting both intestinal and extra-intestinal symptoms up to 4 months following infection.

Different risk factors for infection with Giardia lamblia assemblages A and B in children attending day-care centres.

Giardia lamblia is a major cause of diarrhoea in children, especially those attending day-care centres (DCCs). Only Giardia assemblages A and B infect humans. Given the lack of assemblage-specific epidemiological data, we aimed to identify risk factors for infection by assemblages A and B in DCC attendees. During 2010-2013, 5,015 faecal samples from ≤4-year-old children attending 40 DCCs participating in laboratory surveillance in the Netherlands were tested for Giardia using RT-PCR. Giardia-positive samples were typed for identification of assemblages A and B. We compared child- and DCC-level characteristics of Giardia-positive children with those of Giardia-negative children using mixed-effects logistic regression. Overall, 226 samples (4.5 %) tested positive for Giardia, and assemblages were determined for 138 of them: 62 (45 %) were assemblage A and 76 (55 %) were B. The only risk factor for assemblage A infection was attending DCCs with indoor sandpits and cats during spring/summer (odds ratio [OR] 13.5; 95% CI 1.8-101.3). For assemblage B, risk factors were attending DCCs with dedicated diaper-changing (OR 3.6; 95% CI 1.7-7.6) and laundry (OR 2.3; 95% CI 1.1-4.9) areas. Preventing sick children from attending day-care and having cloth-towels at the DCC decreased the risk of assemblage B infection (OR 0.0; 95% CI 0.0-0.5 and OR 0.3; 95% CI 0.1-0.6 respectively). Risk factors for assemblages A and B infection in DCC-attending children were different, with assemblage B being mainly related to anthroponotic transmission, and assemblage A being related to zoonotic transmission. Given these differences, interventions to reduce the burden of childhood giardiasis cannot ignore those assemblage-specific preferred reservoirs and transmission routes.

Brachyspira species and gastroenteritis in humans.

Brachyspira species have been implicated as a potential cause of gastroenteritis in humans; this is, however, controversial. In 733 gastroenteritis cases and 464 controls, we found 29 samples positive for Brachyspira species (2.3% of cases and 2.6% of controls; P = 0.77). Brachyspira species were not associated with gastroenteritis in humans.

First case of Echinococcus vogeli infection imported to the Netherlands, January 2013.

In January 2013 in the Netherlands, a man in his 50s from Suriname underwent hemihepatectomy because of a cystic liver mass, assumed to be a cystadenoma. Pathology revealed an echinococcal infection. PCR analysis of cyst material identified Echinococcus vogeli, causing polycystic hydatid disease. This echinococcus species is rarely diagnosed outside South America. The patient received adequate treatment, but this case emphasises the importance of awareness of this infection when treating patients with cystic tumours from endemic areas.

Detection and identification of Toxocara canis DNA in bronchoalveolar lavage of infected mice using a novel real-time PCR.

Toxocarosis is a zoonosis with worldwide distribution caused by Toxocara spp. of dogs and cats. In humans, diagnosis relies mainly on detection of parasite-specific antibodies. Although serological assays in current use have defined sensitivity and specificity, the problem of cross-reactivity still remains, particularly in areas of endemic polyparasitism. Microscopic detection of the parasite in tissue biopsies is not recommended for diagnosis because larvae can be difficult to locate, and finding the parasite eggs in faeces is not applicable since the larvae do not develop to the adult stage in the human host. In this study we describe a novel real-time PCR ('Nemo-PCR') that, in combination with DNA sequencing, allows the detection and identification of Toxocara canis and other nematodes in the Superfamily Ascaridoidea. Results indicate that this approach can detect Toxocara spp. DNA in bronchoalveolar lavage (BAL) of experimentally-infected mice. For diagnostic purposes further studies are necessary to evaluate this assay including testing human BAL fluid. The availability of such a direct assay would improve diagnosis of toxocarosis particularly for patients with pulmonary signs and symptoms.

Spontaneous cure of American cutaneous leishmaniasis due to Leishmania naiffi in two Dutch infantry soldiers.

We report two Dutch infantry soldiers who acquired American cutaneous leishmaniasis (ACL) during military jungle training in Surinam. The lesions had existed for 3 and 5 months, respectively, before the soldiers presented for treatment. The lesions occurred on the head and right thigh, and were small, uncomplicated and symptomless. PCR for Leishmania revealed Leishmania naiffi in both patients. No treatment was given, and the lesions in both men healed spontaneously within 4 and 6 weeks, respectively, after presentation to our clinic. CL is one of the important 'tropical' diseases in The Netherlands, primarily due to the increasing numbers of cases in travellers and in military personnel serving overseas. ACL due to L. naiffi is thought to be a mild expression of CL with a self-limiting nature. Lesions seem to be single, mostly small, ulcerating and usually appear on the hands, arms and legs. No case of mucocutaneous leishmaniasis has yet been attributed to this parasite.

Initiation, establishment and maintenance of Hox gene expression patterns in the mouse.

Spatially and temporally restricted expression of the Hox genes along the main and appendicular axes is essential for correct patterning of vertebrate embryos. In this overview we discuss the latest data that shed light on the mechanisms underlying the generation of the expression domains of the Hox genes. The molecular genetic interactions governing initial transcription of the Hox genes in the posterior part of the primitive streak during mouse and chick gastrulation remain enigmatic. But the recent discovery by Kondo and Duboule (Cell, 97, 1999, 407-417) of a "cluster repressive regulation", will undoubtedly lead to a better understanding of the molecular genetic mechanism underlying colinear and sequential initiation of Hox gene transcription. Recently progress has been booked in characterizing the basal processes driving progression of the Hox expression domains during their establishment. Hox expression is still labile while being established. The transcriptional state of Hox genes in anterior tissues can be reprogrammed under the influence of more posterior locations. Posteriorizing activity may involve RA and FGF signaling. It is only when these interactions and, in some cases at least, regulatory interactions with Hox and cdx gene products occur appropriately, that the Hox expression domains would be correctly established. After the Hox expression domains have been established, regulatory processes involving the products of Polycomb and trithorax- Group genes start operating, perpetuating the transcriptional state of the Hox genes within and outside the expression domains. Whether control at the level of chromatin structure, believed to operate during the late maintenance phase of Hox gene expression, is also involved in regulating concerted initial expression of these genes, is a possibility that has been suggested.

Mutation detection in the repeated part of the PKD1 gene.

The principle cause of one of the most prevalent genetic disorders, autosomal dominant polycystic kidney disease, involves mutations in the PKD1 gene. However, since its identification in 1994, only 27 mutations have been published. Detection of mutations has been complicated because the greater part of the gene lies within a genomic region that is reiterated several times at another locus on chromosome 16. Amplification of DNA fragments in the repeated part of the PKD1 gene will lead to coamplification of highly homologous fragments derived from this other locus. These additional fragments severely hamper point-mutation detection. None of the point mutations published to date are located in the repeated part of the PKD1 gene. However, we have reduced the problems posed by the strong homology, by using the protein-truncation test, and we have identified eight novel mutations, seven of which are located in the repeated part of the PKD1 gene.

Detection of translation terminating mutations in the PKD1 gene.

Since the identification of the PKD1 gene in 1994, few mutations have been found. This is mainly due to the very strong homology between a large part of the PKD1 gene and another locus on the short arm of chromosome 16. It is expected that the majority of mutations at the PKD1 locus will be small deletions, insertions and point mutations. Amplification of a piece of DNA derived from the repeated area of the PKD1 gene will result in co-amplification of DNA fragments from the other locus. Detection of mutations is significantly hampered this way. We present a procedure, using the protein truncation test, which reduces the complexity caused by the homologous sequences. We have studied a group of 20 patients with ADPKD at the 3' end of the PKD1 transcript and have produced promising results.

Cytoplasmic acidification facilitates but does not mediate DIF-induced prestalk gene expression in Dictyostelium discoideum.

Stalk cell differentiation in Dictyostelium can be induced by the differentiation-inducing factor, DIF, or by conditions that decrease intracellular pH (pHi). We have investigated whether cytoplasmic acidification acts directly to induce expression of pDd56 and pDd63, two DIF-regulated genes, specifically expressed in prestalk cells. The weak base methylamine, which increases pHi, inhibits DIF-induced transcription. The weak acid 5,5-dimethyl-2,4-oxazolidinedione (DMO), which decreases pHi, stimulates DIF-induction of the two prestalk genes. After relatively long incubation periods, DMO also induces a low level of prestalk gene expression in the absence of added DIF. However, unlike DIF-mediated induction, the apparent DMO-mediated induction decreases to undetectable levels when the cell density is reduced from 10(7) to 10(5) cells/ml. This indicates that DMO does not itself induce gene expression, but acts to enhance the effects of an autonomously secreted stalk-inducing factor, presumably DIF. These results suggest that the effects of DIF on gene expression are regulated by intracellular pH, but do not support a role for protons as direct intermediates in the DIF signal transduction pathway.