Prevalence and Genotype Distribution of Pneumocystis jirovecii in Cuban Infants and Toddlers with Whooping Cough

This study describes the prevalence and genotype distribution of Pneumocystis jirovecii obtained from nasopharyngeal (NP) swabs from immunocompetent Cuban infants and toddlers with whooping cough (WC). A total of 163 NP swabs from 163 young Cuban children with WC who were admitted to the respiratory care units at two pediatric centers were studied. The prevalence of the organism was determined by a quantitative PCR (qPCR) assay targeting the P. jirovecii mitochondrial large subunit (mtLSU) rRNA gene. Genotypes were identified by direct sequencing of mtLSU ribosomal DNA (rDNA) and restriction fragment length polymorphism (RFLP) analysis of the dihydropteroate synthase (DHPS) gene amplicons. qPCR detected P. jirovecii DNA in 48/163 (29.4%) samples. mtLSU rDNA sequence analysis revealed the presence of three different genotypes in the population. Genotype 2 was most common (48%), followed in prevalence by genotypes 1 (23%) and 3 (19%); mixed-genotype infections were seen in 10% of the cases. RFLP analysis of DHPS PCR products revealed four genotypes, 18% of which were associated with resistance to sulfa drugs. Only contact with coughers (prevalence ratio [PR], 3.51 [95% confidence interval {CI}, 1.79 to 6.87]; P = 0.000) and exposure to tobacco smoke (PR, 1.82 [95% CI, 1.14 to 2.92]; P = 0.009) were statistically associated with being colonized by P. jirovecii. The prevalence of P. jirovecii in infants and toddlers with WC and the genotyping results provide evidence that this population represents a potential reservoir and transmission source of P. jirovecii.     

Prevalence and clinical characteristics of human respiratory syncytial virus in Chinese adults with acute respiratory tract infection

Respiratory syncytial virus (RSV) is a leading cause of respiratory tract illnesses worldwide. Although the prevalence and clinical manifestations of the two subtypes, RSV‐A and RSV‐B, have been studied in some detail in infants and young children, they have not been determined in adults. To evaluate the prevalence of the RSV subtypes and disease severity between RSV‐A and RSV‐B infections in adults, nasal and throat swabs that were collected from patients ≥15 years old who sought medical care for acute respiratory infections at the Fever Clinic of the Peking Union Medical College Hospital in Beijing, China between May 2005 and April 2010. The samples were tested for RSV infection using PCR and sequencing analysis. RSV was detected in 95 (1%) of the adult patients, of whom 53 (55.8%) were positive for RSV‐A and 42 (44.2%) for RSV‐B. The incidence of RSV infections increased with age (χ² = 37.17, P = 1.66E?07). Demographic data and clinical manifestations of RSV‐A were similar to those of RSV‐B. Although RSV‐A and RSV‐B co‐circulated during the 2005–2006 and 2008–2009 seasons, RSV‐A was predominant in the 2006–2008 seasons, whereas RSV‐B was predominant in the 2009–2010 season. Upper respiratory tract infections were diagnosed in most RSV‐infected patients (n = 80, 84.2%), and three patients suffered from pulmonary infection. This is the first study to provide data on the prevalence and clinical manifestations of RSV subgroups among Chinese adults with fever and acute illness, over five successive epidemic seasons. J. Med. Virol. 85:348–353, 2013. © 2012 Wiley Periodicals, Inc.     

Helminthiasis and culture change among the Cofán of Ecuador

Blood and fecal samples and lifestyle interviews were obtained in August 1996 from two Cofán villages in Ecuador, Dureno and Zabalo, that are experiencing different degrees of acculturation. Dureno's territory has been divided by roads and encroached upon by oil companies and colonists, whereas Zabalo, 95 km downriver from Dureno, is located in the Cuyabeno Fauna Reserve, a protected area. This study examines how culture change affects the prevalence of parasitic infections. Fecal samples were examined for helminth eggs to determine helminthiasis in each village sample. Seventy-four samples were collected (Dureno n = 50; Zabalo n = 24), representing 20% of the population in each village. Hemoglobin (Hb) levels and eosinophil counts were determined for all adult participants (Dureno n = 39; Zabalo n = 13). Microscopic examination of prepared samples revealed ova from two parasitic nematodes: roundworm (Ascaris lumbercoides) and hookworm (Ancylostoma duodenale or Necator americanus). Roundworm and hookworm infections were higher in Dureno (44% positive) compared to Zabalo (17% positive), with infected adults from both villages showing eosinophilia. Hb levels were not significantly correlated with the prevalence of helminth infections. It appears that the transmission and prevalence of parasitic infections within each community is associated with the interaction of biological, cultural, and environmental factors. Am. J. Hum. Biol. 12:465–477, 2000. © 2000 Wiley-Liss, Inc.     

Prevalence of Dientamoeba fragilis,Giardia duodenalis,Entamoeba histolytica/dispar,and Cryptosporidium spp in Da Nang,Vietnam, detected by a multiplex real‐time PCR

We surveyed the prevalence of Dientamoeba fragilis, Giardia duodenalis, Entamoeba histolytica, Entamoeba dispar, and Cryptosporidium spp in individuals with and without gastrointestinal (GI) symptoms residing in and around Da Nang city, Vietnam. Fecal samples were collected from children (n = 100) and adults (n = 80) with GI symptoms and from healthy individuals (n = 88) reporting no GI symptoms. Parasite detection was performed by multiplex real‐time PCR. Overall, except for G. duodenalis, we found a low prevalence (<5%) of D. fragilis and E. dispar and no detection of E. histolytica and C. spp in all participants with GI symptoms. Specifically for D. fragilis this contrasts with findings in European populations of children with GI symptoms showing prevalence up to 73%. Moreover, our results indicate that the prevalence of G. duodenalis is higher in patients with GI symptoms compared to asymptomatic individuals and this difference is most obvious in young patients.     

Prevalence and genotypes of human parechovirus in stool samples from hospitalized children in Shanghai, China, 2008 and 2009

Human parechoviruses (HPeVs) are widespread pathogens belonging to the Picornavirus family. The aim of the present study was to assess the prevalence and genetic diversity of HPeV in Shanghai, China, during a HPeV screening program in 2008 and 2009. Of 300 stool samples from children under the age of 5 years with acute diarrhea seen at Children's Hospital, Fudan University, Shanghai, China, 165 (55%) were HPeV‐positive. The median age of infected children was 3 months. The prevalence of HPeV was high (57%) in infants up to 2 years old but dropped to 30.4% in children between 2 and 5 years old. The prevalence did not differ by sex. Infections were present throughout the year but peaked in July and August. The most predominant genotype was HPeV1. Of the 139 strains, 4 were found in 9 samples: HPeV4 (n = 4), HPeV5 (n = 1), HPeV6 (n = 1), and HPeV8 (n = 3). This study provided useful data on the epidemiology of HPeV infection by documenting the distribution of genotypes, age of infection, and seasonal patterns in Shanghai, China. J. Med. Virol. 83:1428–1434, 2011. © 2011 Wiley‐Liss, Inc.     

Evaluation of the performance of DiaSorin molecular Pneumocystis jirovecii-CMV multiplex real-time PCR assay from bronchoalveolar lavage samples

The aim of this study was to evaluate the performance of the DiaSorin Molecular PJ-CMV multiplex real-time PCR (PJ-CMV PCR) assay (DiaSorin Molecular LLC, USA) in bronchoalveolar lavage (BAL) samples compared to direct immunofluorescence assay (IFA) for the detection of Pneumocystis jirovecii and assess CMV and P. jirovecii co-infection rate in immunosuppressed patients with suspected pneumonia. A total of 125 BAL samples from immunosuppressed patients submitted for PJP-IFA were tested. Surplus samples were saved and further tested by using the PJ-CMV PCR assay. Among the 125 samples, P. jirovecii was detected in 31.2% (39/125) and in 40% (50/125) of the specimens using IFA and PJ-CMV PCR respectively. Eleven of the PJ-CMV PCR positive samples were negative by direct IFA for P. jirovecii. All samples positive by direct IFA were also positive by PJ-CMV PCR. Using the direct IFA as a gold standard, the PJ-CMV PCR sensitivity, specificity, positive predictive value and negative predictive value for detection of P. jirovecii were 100%, 87.2%, 78% and 100%, respectively. However, after reviewing the clinical diagnosis, the specificity and PPV increased to 100%. Of the 50 P. jirovecii samples positive by PJ-CMV PCR, 18 (36%) were also positive for CMV by the PJ-CMV PCR. The co-infection rate was found to be 37.5% (6/16) and 35.2% (12/34) in HIV infected and non-HIV infected patients. This study indicated that the DiaSorin Molecular PJ-CMV multiplex real-time PCR assay has higher sensitivity than direct IFA for detection of P. jirovecii and provides rapid detection of PJ and CMV infection in BAL samples.     

Review of cytomegalovirus shedding in bodily fluids and relevance to congenital cytomegalovirus infection

Congenital cytomegalovirus (CMV) infections are a leading cause of sensorineural hearing loss (SNHL) and neurological impairment. Congenital transmission of CMV can occur with maternal primary infection, reactivation, or reinfection during pregnancy. We reviewed studies of CMV shedding in bodily fluids (defined as CMV detected by culture or CMV DNA detected by polymerase chain reaction). Following diagnosis at birth, children with congenital CMV infection exhibited the highest prevalences of CMV shedding (median = 80%, number of sample population prevalences [N] = 6) and duration of shedding, with a steep decline by age five. Healthy children attending day care shed more frequently (median = 23%, N = 24) than healthy children not attending day care (median = 12%, N = 11). Peak shedding prevalences in children occurred at 1–2 years of age, confirming that young children are the key transmission risk for pregnant women. CMV shedding among children was more prevalent in urine specimens than in oral secretions (median prevalence difference = 11.5%, N = 12). Adults with risk factors such as STD clinic attendance had higher shedding prevalences (median = 22%, N = 20) than adults without risk factors (median = 7%, N = 44). In adults with risk factors, CMV was shed more frequently in urine; in adults without risk factors genital shedding was most common. The prevalence of CMV shedding in nine sample populations of pregnant women increased with advancing gestation. In seven sample populations of children with congenital CMV infection, higher viral load at birth was consistently associated with an elevated risk of SNHL. Higher CMV viral load at birth also consistently correlated with the presence of symptoms of congenital CMV at birth. Published 2011. This article is a US Government work and is in the public domain in the USA.     

Clinical features and molecular epidemiology of rotavirus and norovirus infections in Libyan children

Rotaviruses and noroviruses are leading viral causes of diarrhoea in children. A cross‐sectional study was undertaken among children aged <5 years with acute gastroenteritis at Al‐Jala Children's Hospital, Tripoli, Libya, from October 2007 to September 2008. Of 1,090 fecal samples collected, 260 from inpatients and 830 from outpatients, all inpatients and approximately a third of outpatients, selected systematically, were investigated for rotavirus and norovirus infection by ELISA and real‐time RT‐PCR, respectively. Of 520 fecal samples examined (inpatients = 260, outpatients = 260), 164 (31.5%) had rotavirus and 91 (17.5%) had norovirus detected. Rotavirus was identified more often among inpatients than outpatients (35.8% vs. 27.3% respectively, P = 0.038). Norovirus was detected more commonly among outpatients than inpatients (21.2% vs. 13.8% respectively, P = 0.028). The peak incidence of infection with both viruses was among children aged between 6 and 11 months. The number of rotavirus cases was highest between November and June with a peak detection rate of 50% in January. Norovirus occurred most commonly from May through August with a peak detection rate of 47% in August. The most prevalent rotavirus genotypes were P[8], G9 (n = 116, 65.9%), followed by P[8],G1 (n = 49, 27.8%); a single P[9], G3 strain was detected. There were seven distinct electropherotypes among the G9 strains and all belonged to VP7 Lineage III. Among 91 noroviruses identified, 90 were genogroup II. Of 26 genogroup II noroviruses examined, all were genotype GII.4. Rotaviruses and noroviruses are both important causes of gastrointestinal infection among young children in Libya. J. Med. Virol. 83:1849–1856, 2011. © 2011 Wiley‐Liss, Inc.     

Adenovirus isolation rates in acute flaccid paralysis patients

Adenoviruses usually cause asymptomatic or mild infection, but occasionally they produce various severe syndromes including neurological disorders. Association of adenovirus infection with acute flaccid paralysis has been investigated. Shedding of adenovirus with feces was detected in 1.05% of young children (mostly infants) with acute flaccid paralysis syndrome versus 0.42% in healthy contact children (P < 0.01). However, 85% of adenoviruses in the pediatric AFP patients belonged to HAdV‐C species, which does not have a known neuropathogenic potential. Also, 40% of adenoviruses were isolated from patients with consequently established diagnosis of traumatic neuritis at the discharge, which was not compatible with virus ethology of neurological lesions. Higher adenovirus prevalence in young neurological patients could be affected by an underlying immune deficiency or by congestion in children's hospitals. Indeed, among 70 patients (40 infants, 30 adults) with primary immune deficiencies, asymptomatic shedding of adenoviruses was found in 10–17%; in one adult patient a mixture of HAdV‐C2 and HAdV‐D15 persisted for several months. Adenoviruses also could be detected in feces of 12% and 57% of healthy young children from two orphanages, respectively. A significant fraction of samples in these groups contained adenovirus mixtures. Therefore, immune deficiencies and congested groups in children's facilities (orphanages and hospitals) could affect significantly the prevalence of adenovirus shedding. The role of adenoviruses in AFP requires further study. J. Med. Virol. 84:75–80, 2011. © 2011 Wiley Periodicals, Inc.     

Seroprevalence of pertussis antitoxin (anti‐PT) in Sweden before and 10 years after the introduction of a universal childhood pertussis vaccination program

Hallander HO, Andersson M, Gustafsson L, Ljungman M, Netterlid E. Seroprevalence of pertussis antitoxin (anti‐PT) in Sweden before and 10 years after the introduction of a universal childhood pertussis vaccination program. APMIS 2009; 117: 912–22. The prevalence of IgG ELISA antibodies against pertussis toxin (anti‐PT) was studied in two Swedish seroepidemiological studies. One was performed in 1997 when the new pertussis vaccination program was 1 year old (n = 3420). In 2007, when Pa vaccines had been used countrywide for 10 years in the universal child vaccination program, this study was repeated to analyze the effect of vaccination on anti‐PT prevalence (n = 2379). Before the statistical analysis of seroprevalence, children vaccinated within the last 2 years before the serosurveys were excluded. The results indicate a reduced exposure to Bordetella pertussis in the population. The proportion of sera without measurable anti‐PT antibodies increased significantly, aggregated over all comparable age groups, from 3.8% in people sampled in 1997 to 16.3% in people sampled in 2007. For cord blood, 1% was without measurable anti‐PT antibodies in 1997 compared to a significantly higher level, 12%, in 2007. With anti‐PT concentrations of ≥50 and ≥100 EU/ml as cutoff points for ‘recent infection’ the proportion above the cutoff points for younger children was significantly higher in 1997 than in 2007 at both cutoff points. For all adults, 20 years of age and older, the difference in proportions above the lower cutoff point was close to statistically significant, comparing 1997 with 2007. This was not the case at 100 EU/ml. In the 1997 samples of children, there was a significant downward trend of ‘recent infections’ at both cutoff points for three sampled age groups between 5 and 15 years of age from 21% at 5.0–5.5 years of age to 7% at 14.7–15.7 years for the lowest cutoff. In the 2007 samples of children, on the contrary, there was a significant continuous upward trend of ‘recent infections’, at both cutoff points, for four sampled age groups between 4 and 18 years of age – from 4% at 4–5 years of age to 16% at 17–18 years at the lowest cutoff. The continuous increase, with age of children with high anti‐PT concentrations, supports the recent change in the general Swedish childhood vaccination program to include a pre‐school booster at 5–6 years and a school‐leaving booster at 14–16 years of age.     

Genome‐wide linkage of obstructive sleep apnoea and high‐density lipoprotein cholesterol in a Filipino family: bivariate linkage analysis of obstructive sleep apnoea

Increasing evidence supports an association between obstructive sleep apnoea (OSA) and metabolic syndrome (MeS) in both children and adults, suggesting a genetic component. However, the genetic relationship between the diseases remains unclear. We performed a bivariate linkage scan on a single Filipino family with a high prevalence of OSA and MeS to explore the genetic pathways underlying these diseases. A large rural family (n = 50, 50% adults) underwent a 10‐cM genome‐wide scan. Fasting blood was used to measure insulin, triglycerides, total cholesterol and high density lipoprotein (HDL) cholesterol. Attended overnight polysomnography was used to quantify the respiratory disturbance index (RDI), a measure of sleep apnoea. Body mass index z‐scores and insulin resistance scores were calculated. Bivariate multipoint linkage analyses were performed on RDI and MeS components. OSA prevalence was 46% (n = 23; nine adults, 14 children) in our participants. MeS phenotype was present in 40% of adults (n = 10) and 48% of children (n = 12). Linkage peaks with a logarithm of odds (LOD) score >3 were demonstrated on chromosome 19q13.4 (LOD = 3.04) for the trait pair RDI and HDL cholesterol. Candidate genes identified in this region include the killer cell immunoglobulin‐like receptor genes. These genes are associated with modulating inflammatory responses in reaction to cellular stress and initiation of atherosclerotic plaque formation. We have identified a novel locus for genetic links between RDI and lipid factors associated with MeS in a chromosomal region containing genes associated with inflammatory responses.     

Homozygous deletion of CDKN2A (p16, p14) and CDKN2B (p15) genes is a poor prognostic factor in adult but not in childhood B-lineage acute lymphoblastic leukemia: a comparative deletion and hypermethylation study

The biological behavior of childhood B-lineage acute lymphoblastic leukemia (B-ALL) is different from that of adults. We performed a comprehensive analysis of the deletion and the methylation profile of CDKN2A (hereafter identified separately as p16 and p14, for the different proteins encoded) and CDKN2B (hereafter p15) in 91 newly diagnosed B-ALL patients (61 children, 30 adults). The prognostic significance of the profiles of these genes and the association between alterations in these genes and known cytogenetic prognostic factors (BCR/ABL; ETV6/RUNX1, formerly TEL/AML1; MLL rearrangement; and ploidy changes of chromosomes) were also assessed. The prevalence of homozygous deletion, hemizygous deletion, and no deletion of the 9p21 region was 11.5%, 16.4%, and 72.1%, respectively, in children and 30.0%, 20.0%, and 50.0%, respectively, in adults; the higher incidence of homozygous deletion in adults was significant (P=0.029). Homozygous deletion was associated with poor overall survival in adults (P=0.019), but not in children. The incidence of promoter methylation of p16, p14, and p15 was 34.4%, 14.8%, and 34.4%, respectively, in children and 26.7%, 10.0%, and 40.0%, respectively, in adults, with no significant difference between the two groups. No significant association was observed between deletion and methylation or with known cytogenetic prognostic factors. The difference in incidence, distribution, and prognostic effect of homozygous deletion in children and adults may explain the prognostic disparity.     

Prevalence of rib anomalies in normal Caucasian children and childhood cancer patients

Purpose. – To evaluate the prevalence of abnormalities of rib development in normal Caucasian children and patients with childhood cancer.Materials And Methods. – Chest radiographs of 881 Caucasian pediatric controls and 906 childhood cancer patients were reviewed, and independently scored by four blinded observers, using strict definitions. Prevalences of 6 major rib anomaly categories in controls were compared to their prevalence in the total group of childhood cancer patients, and the 12 individual larger tumor groups using Chi-square tests.Results. – Values in the control population were generated for the occurrence of six major rib anomaly categories; cervical rib anomalies were present in 6.1% of controls, aplasia of 12^th ribs in 6.6%, lumbar ribs in 0.9%, bifurcations in 0.7%, synostosis-bridging in 0.3%, and segmentations were not found. The overall prevalence of total rib anomalies in cases and controls was equal (14.9% and 14.2%, respectively). Cervical rib anomalies were found significantly more often in cases (8.6%) compared to controls (p-value=0.047), three groups accounting for this higher prevalence: 12.1% of acute lymphoblastic leukemia patients (p=0.011), 18.2% of astrocytoma patients (p=0.023), and 14.7% of germ cell tumor patients (p=0.046) had a cervical rib anomaly.Conclusion. – Prevalence figures for the presence and type of rib anomalies in a large group of normal Caucasian children were generated. In childhood cancer patients a significantly higher prevalence of cervical rib anomalies was demonstrated in patients with acute lymphoblastic leukemia, astrocytoma, and germ cell tumors.     

A new method for the detection of Pneumocystis jirovecii using flow cytometry

Pneumocystis jirovecii is an opportunistic pathogen responsible for severe pneumonia in immunocompromised patients. Its diagnosis has been based upon direct microscopy either by classic staining (Gomori Grocott) or by epifluorescence microscopy (immunofluorescence staining, IFS), both of which are time-consuming and low on sensitivity. Our aim was to develop a flow cytometric (FC) protocol for the detection of P. jirovecii on respiratory samples. In our study, 420 respiratory samples were analysed in parallel by IFS and FC, and compared from clinical diagnosis to its resolution upon specific anti-Pneumocystis therapy. The optimum specific antibody concentration for FC analysis was determined to be 10 μg/ml, without any cross-reactions to bacteria or fungi. All positive cases detected by IFS were positive by FC; however, FC classified eight samples to be positive which were classified as negative by routine technique. These samples were obtained from patients with respiratory symptoms who responded favourably to Pneumocystis-specific therapy and were subsequently considered to be true-positives. Using clinical diagnosis as a reference method, FC showed 100% sensitivity and specificity, whereas IFS showed 90.9% sensitivity and 100% specificity. According to our results, a new diagnostic approach is now available to detect P. jirovecii in respiratory samples.     

Prevalence and cellular reservoir of latent human herpesvirus 6 in tonsillar lymphoid tissue.

There are few studies that examine prevalence, quantity, and cellular proclivity of latent human herpesvirus 6 (HHV-6) in healthy populations. We examined 69 tonsils with paired blood specimens from children without evidence of acute infection. By polymerase chain reaction (PCR), HHV-6 was detected at low levels in 100% of tonsils and 39% of blood samples (n = 27), suggesting that prevalence of latent HHV-6 infection is high in children and may be underestimated by PCR analysis of blood. Although HHV-6A and HHV-6B were detected, HHV-6B predominated, being found in 97% of samples (n = 67). Tonsil sections from 7 cases were examined by in situ hybridization using 2 HHV-6 probes and immunohistochemical analysis. Using both in situ hybridization and immunohistochemical analysis, all tissues revealed marked HHV-6-specific staining in the squamous epithelium of the tonsillar crypts and rare positive lymphocytes. We conclude that HHV-6 is present universally in tonsils of children, and tonsillar epithelium may be an important viral reservoir in latent infection.     

Comparison of Epidemiology,Emergency Care,and Outcomes of Acute Ischemic Stroke between Young Adults and Elderly in Korean Population: A Multicenter Observational Study

Stroke in young adults has been known to show a lower incidence and a better prognosis. Only a few studies have examined the epidemiology and outcomes of ischemic stroke in young adults and compared them with the elderly in Korean population. All consecutive patients with ischemic stroke visiting 29 participating emergency departments were enrolled from November 2007 to October 2009. Patients with less than 15 yr of age and unknown information on age and confirmed diagnosis were excluded. We categorized the patients into young adults (15 to 45 yr) and elderly (46 yr and older) groups. Of 39,156 enrolled all stroke patients, 25,818 with ischemic stroke were included and analyzed (young adult; n=1,431, 5.5%). Young adult patients showed lower prevalence of most chronic diseases but significantly higher prevalence in exercise, current smoking, and alcohol consumption. Hospital mortality was significantly lower in young adults than elderly (1.1% vs. 3.1%, P<0.001). Higher number of patients in elderly group (68.1%) showed worsening change of modified Rankin Scale than young adults (65.2%). Young adults ischemic stroke showed favorable hospital outcomes than the elderly in Korean population.

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Determination of prevalence and risk factors for infection with <Emphasis Type="Italic">Babesia ovis</Emphasis> in small ruminants from Turkey by polymerase chain reaction

In this study, PCR and thin blood smear-based diagnostic methods were used to assess the frequency of Babesia infection in small ruminants. A total of 300 sheep and 100 goats from 37 randomly selected herds located in eight locations of eastern Turkey were examined for the presence of Babesia infection and any tick species on the body of the animals. Of 400 blood samples examined, 6 (1.5%) were positive for Babesia spp. piroplasms upon microscopic examination, whereas 33 (8.25%) were positive for the presence of B. ovis by PCR. The prevalence of babesiosis in small ruminants detected by PCR was significantly higher than obtained in microscopic examination of thin blood smears (P < 0.05). Thirty-three animals produced the DNA fragment specific for Babesia ovis of which 32 (10.66%) were sheep and 1 (1%) was goat. The difference between the prevalence of Babesia infection in sheep and goats were statistically significant (P < 0.05). The prevalence of Babesia infection in different age groups of sheep were statistically non-significant (P > 0.05). The frequency of B. ovis infection was higher in herds with tick burden than no tick burden (P < 0.05). Seven amplicons (six from sheep and one from goat) were sequenced. The resulting sequences were identical to the recently reported nucleotide sequence of B. ovis. A total of 510 ticks belonging to the Rhipicephalus spp. were collected from sheep. Ticks were identified to be R. bursa and R. turanicus on the basis of morphological features. Nucleotide sequences data reported in this paper are available in EMBL, GenBank and DDJB databases under the accession numbers: DQ409332, DQ409333, DQ409334, DQ409335, DQ409336, DQ409337, DQ409338.     

Seasonally of cryptosporidiosis in children

The seasonal distribution of cryptosporidiosis in children in Aragón, a region in northeastern Spain, was determined. Over a period of six years (October 1988 to September 1994), 10,034 stool samples from 4,508 children with gastrointestinal symptoms were analyzed for this purpose. The age of the patients ranged from 1 month to 14 years.Cryptosporidium oocysts were identified in 87 (1.93%) patients. Prevalence was highest (6.20%) in children aged 1 to 3 years old. The prevalence was significantly higher in the autumn-winter period (October to March) than in the spring-summer period (April to September) in the whole population (2.41% vs. 1.35%, p=0.010) and in the 1- to 3-year-old age group (8.44% vs. 3.20%, p=0.002), but not in the other age groups. A possible relationship of this pattern to attendance at child care centres is suggested.     

Alloiococcus otitidis —otitis media pathogen or normal bacterial flora?

During the last decade a new potential otitis media pathogen, Alloiococcus otitidis, has been studied. It is still not clear whether this bacterium really is a pathogen, although it has been found in a high percentage of middle ear effusions in children. The present study aimed to investigate the presence of A. otitidis in the nasopharynx and outer ear canals, and to develop a culture method that would make it possible to isolate A. otitidis from these locations. Nasopharyngeal samples (n=129) from children below 6 years were investigated by conventional culture on blood agar plates with 6% saline and rabbit antisera against A. otitidis, and by a PCR method. In the same way, we investigated 10 samples from vestibulum nasi of healthy persons, 68 samples from outer ear canals of patients with acute or chronic ear problems, and 24 samples from outer ear canals of healthy persons. In a rat model of acute otitis media, we instilled living A. otitidis into rat middle ears through the tympanic bulla and evaluated the outcome clinically by otomicroscopy at days 3, 6 and 14. Of the 129 nasopharyngeal cultures, 9 were positive for A. otitidis by PCR, but none by the culture method. Of the 68 samples from patients with running ears, 4 were positive for A. otitidis by PCR, but none by the culture method. Of the 24 healthy ear canals, 7 were positive for A. otitidis by PCR and 3 of them also by the culture method. No A. otitidis could be found from the vestibulum nasi. The rat experiment showed that the reactions in the middle ears were mild; we could not provoke a purulent acute otitis media in any of the rats. There was a 7% prevalence of A. otitidis in children below 6 years. The highest prevalence (29%) was found in outer ear canals of healthy persons, which strongly suggests that A. otitidis is part of the normal bacterial flora of the outer ear canal. The doubtful pathogenicity is also confirmed by the fact that—in the rat model—A. otitidis elicited only a mild response in the middle ear. It was possible to isolate A. otitidis using a blood agar plate with 6% saline.