Human bocavirus in Jordan: prevalence and clinical symptoms in hospitalised paediatric patients and molecular virus characterisation.

INTRODUCTION: This study was aimed at investigating the prevalence of human bocavirus (HBoV) among Jordanian children hospitalised with lower respiratory tract infection (LRTI) as well as the clinical feature associated with HBoV infection, the seasonal distribution of HBoV and the DNA sequencing of HBoV positive samples. METHODS: A total of 220 nasopharyngeal aspirates were collected from children below 13 years of age who were hospitalised with LRTI in order to detect the presence of HBoV using real-time polymerase chain reaction assay and direct HBoV sequencing. RESULTS: HBoV was detected in 20 (9.1 percent) patients, whose median age was four (range 0.8-12) months. Children under the age of 12 months were more susceptible to HBoV infection (p-value is 0.016). The main clinical diagnoses of patients infected with HBoV were bronchopneumonia (35 percent) and bronchiolitis (30 percent). Coughing (100 percent), wheezing (82.7 percent) and fever (68.2 percent) were the most prominent symptoms in infected patients. HBoV infections were seasonal; increasing in cooler months, diminishing in the summer and peaking in March (45 percent). Direct DNA sequencing revealed that three out of 20 (15 percent) specimens were identical to Stockholm 1 and 2 isolates, and single base pair substitution (A to T) at codon 92 was found in 17 out of the 20 (85 percent) specimens that were positive for HBoV, resulting in a threonine-to-serine substitution. CONCLUSION: More attention should be given to diagnosing HBoV in patients with LRTI using molecular techniques.

Detection of herpes simplex and varicella zoster viruses in clinical specimens using direct immunofluorescence and cell culture assays.

Patients (33 in toto) with a clinical diagnosis of herpes infections (simplex, zoster or chickenpox) were investigated for the presence of herpes simplex virus (HSV) and varicella zoster virus (VZV) in skin samples, using direct immunofluorescence and cell culture assays. Five patients with nonherpetic vesiculobullous disorders were included as negative controls. Of the 33 patients, nineteen (57.6%) were positive for HSV or VZV and fourteen (42.4%) were negative. Five controls were all negative for HSV or VZV. Of the nineteen positive patients, HSV was isolated from eight (42.1%) patients, by both direct immunofluorescence and cell culture assays. VZV was isolated from eleven (57.9%) patients, eleven (100%) by direct immunofluorescence assay, and six (54.5%) by cell culture assays. HSV was isolated from one patient clinically diagnosed as chickenpox (VZV), but otherwise the positive laboratory results were concordant with the clinical diagnosis. For epidemiological studies, atypical cases and immunocompromised patients the clinical diagnosis should be confirmed in the laboratory.

Detection of adenovirus infection in children in Jordan.

Between November 1997 and May 1998, 350 nasopharyngeal aspirates (NPA) were obtained from children admitted to the Respiratory Disease Unit at Princess Rahma Hospital, northern Jordan who were clinically diagnosed as suffering from respiratory tract infections. NPA were investigated for the presence of adenovirus using shell vial (SV) culture assay, conventional culture (CC) assay, and direct immunofluorescence assay (DFA). Of the 350 NPA, adenoviruses were detected in 54 (15.4%) by the combined techniques used. SV identified 34 (63%), CC 48 (89%) and DFA 30 (56%). Most virus isolations were in children aged 1-< 5 years old and were associated with pneumonia in 39% and bronchopneumonia in 32%. SV assay showed a sensitivity and specificity of 68.8% and 99.7%, respectively, for detecting adenovirus from NPA. These results emphasize that CC assay is still important for the diagnosis of adenovirus, although SV and DFA are superior diagnostic assays.

Bacteremia in children: etiologic agents, focal sites, and risk factors.

A prospective study was carried out on 210 cases of children under 10 years of age with fever. Cases of gastroenteritis, respiratory tract infections, and suspected sepsis in children seen or admitted to the pediatric hospital were studied. Clinical and microbiological data were recorded in a questionnaire or obtained from patient medical records. Most of the children with septicemia (71.3 per cent) were less than 1 year old. Focal source of bacteremia was gastroenteritis (40.4 per cent), pneumonia or bronchopneumonia (20 per cent), meningitis (7.4 per cent), and urinary tract infections (7.4 per cent). The predominant pathogens isolated from blood or stool specimens were gram-positive bacteria (53.3 per cent), mainly Streptococcus pneumoniae and coagulase-negative Staphylococcus spp. The gram-negative bacteria (45.6 per cent) were mainly Escherichia coli, Klebsiella pneumoniae, Haemophilus influenzae, Neisseria meningitidis, and Yersinia spp. One case of Candida albicans (1.1 per cent) was reported. Pasteurella pneumotropica was reported in two cases for the first time. The mortality rate was 4 per cent, mostly from septicemia cases. Long duration of hospitalization (> 10 days) and parenteral feeding were identified as risk factors. Resistance of the isolated pathogens to several commonly used antibiotics was observed. Empirical treatment with antibiotics is recommended only in life-threatening cases.

Enhanced detection of respiratory syncytial virus by shell vial in children hospitalised with respiratory illnesses in northern Jordan.

During the period between November 1997 and May 1998, a total of 350 nasopharyngeal aspirates were obtained from children admitted to the Respiratory Disease Unit at Princess Rahma Hospital, northern Jordan, and diagnosed clinically as suffering from respiratory tract infections. Nasopharyngeal aspirates were investigated for the presence of respiratory syncycial virus (RSV) by using shell vial (SV) culture assay, conventional culture assay, and direct immunofluorescence assay. Out of 350 nasopharyngeal aspirates, 101(28.9%) were found positive by any of SV, conventional culture, and immunofluorescence; 91 (90.1%) by SV, 87(86.1%) by culture, and 83(82.2%) by immunofluorescence. The maximum number of virus isolations was noted in children below the age of 1 year and was associated with bronchiolitis. SV assay showed the highest sensitivity (94.3%) and specificity (96.9%) for detecting RSV from nasopharyngeal aspirates. These results emphasise the importance of SV culture assay for diagnosis of RSV, although immunofluorescence is a valuable, rapid diagnostic assay.

Seroepidemiologic study of herpes simplex virus type 2 and cytomegalovirus among young adults in northern Jordan.

Blood samples were randomly collected from 360 males and 390 females among apparently healthy university students aged 18-24 years and tested for herpes virus type 2 (HSV-2) and cytomegalovirus (CMV) antibodies. The prevalence of HSV-2 seropositivity was 52.8% for males and 41.5% for females as detected by ELISA. On the other hand, the prevalence of CMV seropositivity was 75.6% in males and 77.2% in females. The high percentage of seropositivity in our study is most probably due to the crowded living and low socioeconomic status of the Jordan population. The higher prevalence in males could be due to the way of life in the Middle East that gives males the freedom to play outdoors more than females.

Prevalence of parainfluenza and influenza viruses amongst children with upper respiratory tract infection.

OBJECTIVE: The objective of this study was to determine the prevalence of Parainfluenza and Influenza causing upper respiratory tract infections and to evaluate shell vial culture assay and direct immunofluorescence assay. METHODS: A retrospective study during the period between November 1997 and May 1998. A total of 350 nasopharyngeal aspirates were obtained from children suffering from respiratory tract infections. Nasopharyngeal aspirates were investigated for the presence of Parainfluenza 1, 2 and 3, Influenza A and B using shell vial culture assay, conventional culture assay and direct immunofluorescence assay. RESULTS: Parainfluenza 1 were identified in 3%, Parainfluenza 2 in 5% and Parainfluenza 3 in 6%. Influenza A were identified in 4% and Influenza B in 2%. Parainfluenza 1, 2 and 3 were isolated in children less than 5 years old. Most of Parainfluenza cases were associated with other upper respiratory infections. Shell vial assay showed a sensitivity of 90-93% and specificity of 99-100% for detecting Parainfluenza 1, 2 and 3. CONCLUSION: These results emphasize that shell vial assay is important for the diagnosis of Parainfluenza and Influenza, although direct immunofluorescence assay is the superior diagnostic assay.

Detection of genetic polymorphism by RAPD-PCR among isolates of Bacillus thuringiensis.

Sixteen isolates of Bacillus thuringiensis recovered from different Jordanian habitats were compared using random amplification of polymorphic DNA (RAPD) to determine whether they could be differentiated at the molecular level. Total genomic DNA from each isolate and three reference strains were amplified using 10-mer primers. Electrophoretic analysis of the amplification products revealed the incidence of polymorphism among the isolates. Pair-wise comparisons of polymorphic products were used to construct a dendrogram applying the cluster analysis. Fifteen of the isolates were all in one major cluster which was divided into six small groups. Such analysis showed some regional variation among the isolates, but did not indicate a clearly defined habitat locational pattern of the DNA polymorphism.

Use of RAPD-PCR fingerprinting to detect genetic diversity of soil Streptomyces isolates.

Random amplified polymorphic DNA (RAPD) was used for identification and assessment of genetic diversity between isolates of Streptomyces from soil. Genomic DNA from 18 Streptomyces isolates and 2 reference strains were amplified using four different 10-mer primers. Different DNA fingerprinting patterns were obtained for all the isolates. Electrophoretic and cluster analysis of the amplification products revealed incidence of polymorphism among the isolates and none of them was identical to the reference strains although there were some common amplification bands. Two highly divergent groups were determined among the isolates. The results indicate that RAPD is an efficient method for discriminating and studying genetic diversity of Streptomyces isolates.

Respiratory syncytial virus infection in infants hospitalized with respiratory illness in northern Jordan.

During the winter seasons of 1993 and 1994, a total of 256 nasopharyngeal aspirates (NPA) from infants aged less than 1 year old admitted to the pediatric ward of Princess Rahma Hospital, northern Jordan, with bronchiolitis and/or pneumonia, were tested for the presence of respiratory syncytial virus (RSV) using direct immunofluorescence assay (DFA) and cell culture (CC). Of the 256 specimens, 129 (50 per cent) were found positive by both DFA and CC, whereas 24 specimens (9 per cent) and 16 specimens (6 per cent) were found positive by DFA and CC, respectively. In an evaluation of the collected NPA specimens detected by DFA, a sensitivity of 89 per cent and a specificity of 78 per cent were demonstrated. These data suggest that virus isolation in CC is still important for the diagnosis of RSV, although DFA is a valuable, rapid diagnostic assay.

Larvicidal activity of Bacillus thuringiensis isolated from Jordanian habitats against Drosophila melanogaster larvae.

Bacillus thuringiensis was isolated from 23 of 37 samples obtained from different Jordanian habitats. Of the 37 samples, 187 different spore-forming colonies were selected and thirty (16%) were identified as B. thuringiensis based on their pathogenicity and production of parasporal inclusions. The lethal dose (LD50) of B. thuringiensis indicated a variation in their pathogenicity to Drosophila melanogaster larvae. Sensitivity of the first, the second and the third instar larvae of D. melanogaster showed slight variation in between. The third instar was the most sensitive stage to the pathogen spores.

Non-seasonal viral and bacterial episode of diarrhoea in the Jordan Valley, West of Jordan.

A non-seasonal diarrhoeal episode in the Jordan Valley occurred over a 2-month period, during which no traditional enteropathogens were detected by the health authority laboratories. A total of 17 diarrhoeal stool specimens from infants, young children and adults were randomly collected and delivered to our laboratories to investigate the presence of unusual aetiological agents. Stools were examined for parasites, ova, viruses and cultured for bacterial pathogens. A multiplex polymerase chain reaction was developed to investigate the involvement of diarrhoeagenic Escherichia coli in this episode. Recognised pathogenic organisms were detected in 8 out of 17 of the diarrhoeatic patients, one patient of whom had a mixed infection with two agents. Rotavirus, enteroinvasive E. coli (EIEC), enteropathogenic E. coli (EPEC), and enterotoxigenic E. coli (ETEC) were found to be associated with the diarrhoea. EIEC was the most common enteropathogen detected (4 out of 17) followed by rotavirus (3 out of 17). One of the EIEC isolates detected in one patient was associated with rotavirus. The clinical features of the diarrhoeatic patients were remarkably similar, regardless of aetiology. This study reveals the identity of pathogenic agents that are not detected by traditional methods employed by the health authority laboratories, which emphasise the urgent need for developing the current diagnostic techniques.

Isolation and phenotypical identification of spore-forming Bacillus species from Jordanian habitats with larvicidal activity against Drosophila melanogaster.

Spore-forming Bacillus isolates were recovered from different Jordanian habitats. Of 37 samples, 187 colonies were selected. Forty-six (24.6%) of them were pathogenic to the third instar larvae of Drosophila melanogaster. Larvicidal activity of the isolates was from 0% (non-cultivated soil) to 83.3% (decomposed animal residues). The total spore count per gram weight varied from 0.1 x 10(5)-18 x 10(5) among the 37 tested samples. Morphological and microscopical identification of the isolates showed the presence of 16 different Bacillus species. The pathogenic isolates were B. thuringiensis (44) and B. sphaericus (2).

Viral gastroenteritis among young children in northern Jordan.

During the summer months of 1992 and 1993, a total of 439 diarrhoeatic fecal specimens from infants and young children less than 3 years of age admitted to the pediatric ward of Princess Basma Teaching Hospital, northern Jordan were tested for the presence of viruses using direct electron microscopy (EM) and enzyme-linked immunosorbent assay (ELISA) for rotavirus. EM revealed rotaviruses in 83 (18.9 per cent) of cases, adenoviruses in five (1.1 per cent) cases, and small round viruses in three (0.68 per cent) cases. In contrast, the ELISA assay detected rotaviruses in 174 (39.6 per cent) of cases. In an evaluation of the collected diarrhoeatic fecal samples for rotavirus detected by ELISA, a sensitivity of 95.2 per cent and a specificity of 73.3 per cent was demonstrated.