Molecular characterization of Echinococcus granulosus in a hyperendemic European focus,the Republic of Moldova

Cystic echinococcosis is a zoonosis caused by the tapeworm Echinococcus granulosus sensu lato. The lifecycle of the parasite is mainly domestic, requiring dogs as definitive hosts and livestock species as intermediate hosts. Although human cystic echinococcosis is a high public health priority in the Republic of Moldova, the rare animal data available concerns only infection in cattle. A preliminary slaughterhouse survey was conducted to assess prevalence and perform the first molecular characterization of E. granulosus sensu lato in sheep and cattle. For the survey, 40 sheep and 19 cattle were inspected. Very high prevalence in sheep (82.5 %) and in cattle (78.9 %) was found. Molecular analyses identified genotypes G1 and G3 of E. granulosus sensu stricto in all the liver and lung samples. Based on the concatenated sequences of cox1?+?nad3 (701 bp), 23 different haplotypes were obtained. Mixed infections by different haplotypes/genotypes were frequently identified in both sheep and cattle. The relatively high (20.0 %) cyst fertility observed in cattle argues for the potential contribution of cattle to the lifecycle of E. granulosus sensu stricto, unlike previous observations in Europe. The hyperendemic situation of Moldova can be explained by a high majority of animals slaughtered at home usually without veterinary inspection. Further extensive slaughterhouse surveys with molecular identification also involving pigs and goats are needed to obtain a better overview of the epidemiological situation of E. granulosus sensu lato in this hyperendemic focus in the Republic of Moldova.     

Taxonomic position and geographical distribution of the common sheep G1 and camel G6 strains of <Emphasis Type="Italic">Echinococcus granulosus</Emphasis> in three African countries

The taxonomic and phylogenetic status of Echinococcus granulosus strains are still controversial and under discussion. In the present study, we investigated the genetic polymorphism of E. granulosus isolates originating from three countries of Africa, including a region of Algeria, where the common G1 sheep and the camel G6 strains coexist sympatrically. Seventy-one hydatid cysts were collected from sheep, cattle, camels, and humans. Two mitochondrial markers (cox1 and nad1) were used for strain identification. Two nuclear markers (actII and hbx2) were used to study the possible occurrence of cross-fertilization. Despite the heterogeneity observed among the G1 isolates, they were all localized within one robust cluster. A second strong cluster was also observed containing all of the G6 isolates. Both strains appeared as two distinct groups, and no cases of interbreeding were found. Thus, the attribution of a species rank can be suggested. We also found the Tasmanian sheep G2 strain for the first time in Africa. Because of the slight variations observed between the common sheep and the Tasmanian sheep strains, further studies should be carried out to elucidate the epidemiological relevance of this genetic discrimination.     

Prevalence and diversity of cystic echinococcosis in livestock in Maasailand, Kenya

Cystic echinococcosis (CE) is a zoonotic disease caused by several members of the Echinococcus granulosus species complex. In East Africa, several species/strains are known to occur in livestock and humans, but host preferences, relative frequencies and spatial distribution of these taxa are poorly known. Here, we contribute livestock data for Maasailand of southern Kenya. Total CE prevalence was 25.8?% in cattle (151/587), 16.5?% in sheep (71/430) and 10.8?% in goats (21/194), which is a significant increase compared to surveys done about three decades ago. The majority of cysts occurred in the liver (56?% in cattle, 70?% in sheep and 65?% in goats). Molecular characterization by PCR?CRFLP and sequencing of parts of the mitochondrial nad-1 gene was done for a subsample of 285 cysts. E. granulosus G1 was dominant in all host species (200 of 201 cysts from cattle, 68 of 69 from sheep and 11 of 15 from goats); the remaining taxa were Echinococcus canadensis G6 (one cyst from sheep, four from goats) and Echinococcus ortleppi (one cyst from cattle). Considering cyst fertility, sheep appear to be the most important hosts for E. granulosus G1, while goats were found to be suitable hosts for E. canadensis G6 (three of four cysts were fertile). For the first time, E. ortleppi was found in cattle from southern Kenya. Our data show an intense and possibly increasing level of CE transmission in southern Kenya, and the predominance of E. granulosus G1, which appears to be particularly pathogenic to humans, calls for urgent control measures.     

Genetic variations among Echinococcus granulosus isolates in Egypt using RAPD-PCR

Cystic echinococcosis (CE), caused by hydatid cysts, is a widespread and hazardous disease in humans and animals worldwide. The aim of the current study was to investigate the genetic variations among Echinococcus granulosus cyst strains isolated from sheep, camel, pig, and donkey using RAPD-PCR analysis. Seven primers of arbitrary sequences were used in the PCR reactions. The screened primers gave total patterns ranging from 27 to 39 reproducible bands for each isolate. Each population isolate gave its specific pattern. Although distinct polymorphic patterns were obtained among the four isolates, there were several shared bands among them in each primer used. A comparison of the different RAPD-PCR patterns showed that primers P1, P3, and OPH 04 yielded band patterns that revealed a high degree of divergence among the four isolates of E. granulosus that allowed easy distinction between them. The remaining primers (P2, P4, P5, and OPH14) amplified DNA fragments that were common to two or more isolates but diversified in the other two or three isolates. The study revealed that the most closely related isolates were of donkey and camel where the similarity coefficent between them ranging from 53?% to 78?%, followed by isolates of pig and sheep (sc?=?40?% to 68?%), while the similarity coefficent between isolates of camel and sheep was 33–45?%, between camel and pig was 36 to 57?%, between donkey and pig was 37 to 52?%, and between donkey and sheep was 35 to 54?% which means that they more or distant from each other. In conclusion, hydatid cysts isolated from camel may have the genotypic characters of donkey strain.     

Copro-DNA test for diagnosis of canine echinococcosis

Echinococcosis is one of the most important parasitic zoonotic diseases in the world. The life cycle of Echinococcus granulosus is dependent to the dog–sheep cycle and is actively transmitted in all pastoral regions where sheep, cattle and camels predominate. DNA approaches are now being used routinely for accurate identification of Echinococcus and Taenia species, subspecies and strains. In this study, faecal samples were collected from 50 stray dogs from Mashhad city in the northeast of Iran during June 2011. All samples were frozen at least for 1 week in ?80°C. The embryophore layer of the eggs was broken by freezing–thawing method after egg concentration by the formalin-ether method. DNA was extracted using a DNA isolation kit (MBST, Germany/Iran) according to the manufacturer’s instructions. After PCR, by the primers expressed in the following it became, clear that about 20 % of stray dogs are infected with E. granulosus. In this study, we describe a modified method for DNA isolation from faeces for coprodiagnosis of Echinococcus spp.     

Echinococcus granulosus of camel origin: development in dogs and parasite morphology

The developmental characteristics ofEchinococcus granulosus of camel origin were studied in four dogs artificially infected with protoscolices originating from hydatid cysts isolated from the lung of a camel (Camelus dromedarius). Two dogs each were necropsied 34/35 and 41 days post-infection (p.i.); one dog had a low worm burden and the others were heavily infected (27,625–41,150 worms). At day 35 p.i., 20% of the parasites had developed three segments and the uterus of the vast majority of the total population was full of developing eggs in the terminal segment. At day 41, up to 58% of the parasites contained mature eggs (embryonated eggs with fully developed, thick-shelled embryophores). Morphological studies revealed the following major characteristics for 35 day-old worms: the mean length of the terminal segment accounted for 54% of the total worm length; the position of the sexually mature segment was always terminal; the female reproductive system possessed an elongated ovary with compact lobules; the female ducts were also compact; the Mehlis' gland was covered by the vitelline gland and the testes were distributed throughout the segment, with 1 row posterior to the vitelline gland. The camel isolate can readily be distinguished from the horse and sheep strains, but it is similar to the cattle strain in some respects, particularly in its precocious development. However, the camel isolate differs from the cattle strain in the position of the sexually mature segment, arrangement of the testes and structure of the female reproductive system. As in the cattle strain, the metacestodes in the principal intermediate host are mostly localised in the lung and have a high fertility rate. Detailed morphological characteristics and biological features of the sheep, cattle, horse and camel isolates ofE. granulosus are presented.Increasing evidence is being accumulated that local populations ofEchinococcus granulosus may vary in morphological and biological features and that the occurrence of intraspecific variants or strains can play an important role in the epidemiology of echinococcosis (Rausch 1986; McManus and Smyth 1986; Thompson 1986; Thompson 1988; Thompson and Allsopp 1988; Eckert and Thompson 1988), because strains differ in their infectivity to final and intermediate hosts and their transmission patterns. Moreover, the pathogenic significance of strains to humans may also vary.The camel is an important host ofE. granulosus and is commonly infected throughout Africa and the Middle East (Schwabe 1986). The camel has attracted much interest as an intermediate host ofE. granulosus, particularly in its role as a reservoir of infection in humans. In addition, it has been proposed that the camel form ofE. granulosus may be a different subspecies or strain (Pandey et al. 1986). However, very little comparative work has been carried out to characterise isolates ofE. granulosus from camels (Thompson and Lymbery 1988).The aim of the present work was to study the morphological and biological characteristics of intestinal stages ofE. granulosus of camel origin obtained from artificially infected dogs. In addition, data were collected on the metacestodal stage.     

Cystic echinococcosis in Turkey: genetic variability and first record of the pig strain (G7) in the country

A sample of 22 Echinococcus granulosus isolates collected from 12 sheep and ten humans from a focus of cystic echinococcosis in western Turkey was examined by DNA sequencing of four mitochondrial genes (cox1, atp6, nad1, rrnS). Results demonstrated the presence of two species of E. granulosus complex, E. granulosus sensu stricto and E. canadensis. Of E. granulosus sensu stricto, the G1 genotype (including three microvariants) was found in 17 isolates from humans and sheep, the G3 genotype and an intermediate form G1/G3 in one isolate each (both from sheep). Of E. canadensis, the pig strain G7 was found in three isolates from sheep and human. This is the first report of this strain in Turkey. Its presence has implications for local control programs due to its shorter maturation rate in dogs compared with E. granulosus sensu stricto. Goat and/or wild boar are likely reservoirs for G7 in the region. We provided further data on the pattern and frequency of nucleotide substitutions within the G1/G3 cluster. Based on our results and GenBank records, G2 (Tasmanian sheep strain) is not considered as a discrete genotypic unit, as its sequences at polymorphic sites conform to microvariants of both G1 and (more often) G3.     

The phylogenetic similarity of hydatid cyst isolated from humans and sheep in Ilam Province southwest of Iran

Hydatid cyst is a chronic zoonotic disease caused by the larval stage of the dog tapeworm, Echinococcus granulosus. To identify genotype of hydatid cysts of human and sheep jackal in Ilam Province (South West of Iran), the PCR-RFLP and DNA sequencing were used. A total of 10 human and 20 sheep protoscoleces hydatid cyst samples were collected from different hospitals and slaughterhouses. Then, the gene of cox1 of mitDNA of the parasite was amplified and PCR products were cut using AluI and HpaII restriction enzymes. Finally, a number of PCR products were bi-directionally sequenced. Based on the DNA sequencing and PCR-RFLP results, human and sheep samples indicated to pertain the genotypic similarities. Our data indicated that, the genotypes of larval stage of E. granulosus is similar in both intermediate hosts. According to the phylogenetic tree, there is at least one genotype of parasite, which belongs to E. granulosous sensu stricto (G1–G3) complex and overall isolates sequences of mtDNA indicated 100 % homology with references G1, G2, and G3 sequences in the GenBank database. G1 genotype was the dominant genotype of human and livestock.     

Molecular characterization of Echinococcus granulosus strains in Sardinia

Investigations were undertaken to determine the genotypes of the parasite Echinococcus granulosus that were present in livestock animals on the island of Sardinia. Liver, lung, and spleen samples were obtained from 770 sheep, 229 cattle, and 277 pigs slaughtered in Sardinia between January 2003 and April 2005, and the number and fertility of hydatid cysts were determined. Protoscoleces and/or germinal layer were collected from individual cysts, DNA was extracted from 91 samples, and polymerase chain reaction (PCR) and PCR-restriction fragment length polymorphism (RFLP) methods were used for identification of the strain genotype for each sample (G1, G5, G6/G7). Fragments of the mitochondrial cytochrome c oxidase subunit 1 and NADH dehydrogenase I were sequenced. Hydatid disease prevalence of 75.3, 41.5, and 9.4% were found in the organs collected from sheep, cattle, and pigs, respectively. Molecular analysis showed that 89 of 91 ovine, bovine, and swine cysts belonged to the G1 genotype (common sheep strain) of E. granulosus. Parasite isolates from two pigs were identified to belong to the G7 genotype (pig strain). Our results confirm the high prevalence of E. granulosus infection in livestock animals in Sardinia and reveal the presence of at least two parasite genotypes in Sardinia.     

Echinococcus spp. in central Kenya: a different story

Research on cystic echinococcosis (CE) has a long history in Kenya, but has mainly concentrated on two discrete areas, Turkana and Maasailand, which are known to be foci of human CE in Africa. Here, we report on a survey for CE in livestock from central to northeastern Kenya, from where no previous data are available. A total of 7,831 livestock carcasses were surveyed. CE prevalence was 1.92 % in cattle (n?=?4,595), 6.94 % in camels (n?=?216), 0.37 % in goats (n?=?2,955) and 4.62 % in sheep (n?=?65). Identification of the parasite was done using an RFLP-PCR of the mitochondrial nad1 gene, which had been validated before against the various Echinococcus taxa currently recognized as distinct species. From a total of 284 recovered cysts, 258 could be identified as Echinococcus granulosus sensu stricto (n?=?160), E. ortleppi (n?=?51) and E. canadensis (n?=?47) by RFLP-PCR of nad1. In cattle, fertile cysts occurred mostly in the lungs and belonged to E. ortleppi (31 of 54), while the vast majority were sterile or calcified cysts of E. granulosus s.s.. Most fertile cysts in camels belonged to E. canadensis (33 of 37); sterile or calcified cysts were rare. Goats harboured fertile cysts of E. ortleppi (n?=?3)—which is the first record in that host species—and E. canadensis (n?=?1), while all cysts of E. granulosus were sterile. Only sterile cysts were found in the three examined sheep. Typically, all cysts in animals with multiple infections belonged to the same species, while mixed infections were rare. Our data indicate that the epidemiological situation in central to northeastern Kenya is clearly different from the well-studied pastoral regions of Turkana and Maasailand, and the apparently low number of human CE cases correlates with the infrequent occurrence of E. granulosus s.s.     

Development of a real-time PCR for the differentiation of the G1 and G2/G3 genotypes of Echinococcus granulosus

The present study was aimed at developing a SYBR Green™-based real-time polymerase chain reaction (PCR) assay for a rapid differentiation of the genotype G1 from the cluster of genotypes G2/G3 of Echinococcus granulosus, using as marker the 12S mtDNA gene. Eleven hydatid cysts from water buffaloes and 19 from cattle were used. Fourteen samples (identified as G1 using sequencing) showed a mean melting temperature (T _m) of 76.4°C and 16 samples (identified as G2/G3 using sequencing) showed a mean T _m of 77.0°C. The detected mean difference of the T _m of 0.6°C between G1 and G2/G3 genotypes might allow a fast and simple discrimination of these genotypes. In conclusion, the real-time PCR developed in the present study provides a powerful tool for molecular studies on E. granulosus with possibilities for extension to other genotypes using different molecular targets. Maria Paola Maurelli and Laura Rinaldi contributed equally to this work.     

An immunoblot for detection of Taenia saginata cysticercosis

Control measures to prevent human infections with the food-borne zoonotic helminth Taenia saginata are currently based on meat inspection, which shows rather low diagnostic sensitivity. To develop an immunoblot for detection of T. saginata-infected cattle, crude proteins of T. saginata cysts were extracted and separated with SDS-PAGE. The cyst antigens showed ten protein bands ranging from 260 to 14 kDa. T. saginata cyst proteins 260, 150, 130, 67, 60, 55, 50, and 23 kDa were immunoreactive with known positive sera of T. saginata-infected cattle but cross-reacted with sera from Echinocccus granulosus-infected ruminants. By contrast, 14- and 18-kDa cyst proteins reacted specifically with T. saginata-positive sera and thus might be potential candidates for the development of a T. saginata-specific immunoassay. Proteins of E. granulosus cysts and Taenia hydatigena cysts were also extracted and separated with SDS-PAGE. E. granulosus cysts revealed 11 protein bands ranging from 260 to 23 kDa. E. granulosus protein 60 kDa was immunoreactive with E. granulosus-positive sera only. The cyst of T. hydatigena showed 11 protein bands ranging from 290 to 14 kDa. The protein band 35 kDa showed cross-reaction with positive sera from both T. saginata- and E. granulosus-infected animals. A protein of 67 kDa was present in all three tested cestode species and was the major antigenic protein detected by sera of T. saginata- and E. granulosus-infected animals. Therefore, this protein represents a potential vaccine candidate against both cysticercosis and cystic echinococcosis in cattle.     

Prevalence survey and first molecular characterization of Echinococcus granulosus in France

Human cystic echinococcosis (hydatid disease) caused by the Echinococcus granulosus tapeworm continues to be a substantial cause of morbidity and mortality in many parts of the world. France is still considered as endemic area, but the current infestation by E. granulosus of intermediate hosts in France remains currently unknown due to the absence of official data reporting for the last 20 years. A 1-year prevalence survey was conducted in the 24 slaughterhouses of ten departments of the South of France. We demonstrate that the E. granulosus parasite is still currently present at low prevalence at slaughterhouses in the study area (4 cases for 100,000 sheep and 3 cases for 100,000 cattle). In addition, we assess the presence of genotype G1 in infected animals and identify for the first time in France genotypes G2 and G3 of E. granulosus sensu stricto.     

Sequence analysis of cox1 and nad1 genes in Echinococcus granulosus G3 genotype in camels (Camelus dromedarius) from central Iran

Nineteen hydatid cyst isolates collected from camels in central Iran were subjected to sequences analysis of mitochondrial cytochrome c oxidase subunit 1 (cox1) and NADH dehydrogenase subunit 1 (nad1) genes. A consensus sequence obtained containing 366 nucleotides for cox1 and 471 nucleotides for nad1 genes. Overall, the camel isolates indicated five different sequences in cox1 and nine in nad1 genes. The sequences analysis indicated that 26.3%, 42.1%, and 31.6% of isolates belonging to G1, G3, and G6 genotypes of Echinococcus granulosus, respectively. The isolates with G3 genotype indicated one cox1 sequence having 100% homology with reference G3 sequence (AN: M84663) and two different nad1 sequences, one having100% homology with reference G3 sequence (AN: AJ237634) and the other with a silent mutation (G to A) in position 279. The presence of G3 genotype (buffalo strain) of E. granulosus as dominant genotype in camels is emphasized. As G3 genotype has formerly been reported in human, the epidemiological role of camels is warranted in future surveys.     

Shiga toxin-producing Escherichia coli in Central Greece: prevalence and virulence genes of O157:H7 and non-O157 in animal feces, vegetables, and humans

In Greece, Shiga toxin-producing Escherichia coli (STEC) have only been sporadically reported. The objective of this study was to estimate the prevalence of STEC and Escherichia coli O157:H7 in farm animals, vegetables, and humans in Greece. A total number of 1,010 fecal samples were collected from farm animals (sheep, goats, cattle, chickens, pigs), 667 diarrheal samples from humans, and 60 from vegetables, which were cultured in specific media for STEC isolates. Enzyme-linked immunosorbent assay (ELISA) was used to detect toxin-producing colonies, which, subsequently, were subjected to a multiplex polymerase chain reaction (PCR) for stx1, stx2, eae, rfbE _O157, and fliC _h7 genes. Eighty isolates (7.9 %) from animal samples were found to produce Shiga toxin by ELISA, while by PCR, O157 STEC isolates were detected from 8 (0.8 %) samples and non-O157 STEC isolates from 43 (4.2 %) samples. STEC isolates were recovered mainly from sheep and goats, rarely from cattle, and not from pigs and chickens, suggesting that small ruminants constitute a potential risk for human infections. However, only three human specimens (0.4 %) were positive for the detection of Shiga toxins and all were PCR-negative. Similarly, all 60 vegetable samples were negative for toxin production and for toxin genes, but three samples (two roman rockets and one spinach) were positive by PCR for rfbE _O157 and fliC _h7 genes. These findings indicate that sheep, goats, cattle, and leafy vegetables can be a reservoir of STEC and Escherichia coli O157:H7 isolates in Greece, which are still rarely detected among humans.     

<Emphasis Type="Italic">Echinococcus granulosus</Emphasis> strain typing in Bulgaria: the G1 genotype is predominant in intermediate and definitive wild hosts

Addressing the genetic variability in Echinococcus granulosus is epidemiologically important, as strain characteristics may influence the local transmission patterns of zoonotic cystic echinococcosis. To classify the genotype(s) present in intermediate (pig, cattle and sheep) and definitive (jackal and wolf) hosts in Bulgaria, a DNA-based approach was used to assess parasite protoscoleces or strobiles. Genes corresponding to coding and non-coding regions of the nuclear and mitochondrial genome (ND-1, HBX, Act II, AgB-1) were amplified by PCR and subsequently sequenced. The sequences resolved were all found to be identical to those published for the common sheep strain of E. granulosus, indicating that the G1 genotype is predominant in Bulgaria. One microvariant for ND-1 was found in the pig isolates; however no epidemiological significance was attributed to this finding.     

Epidemiological studies on Echinococcus in Pampas fox (Lycalopex gymnocercus) and European hare (Lepus europaeus) in Buenos Aires province, Argentina

In Argentina, hydatid disease caused by Echinococcus granulosus is widespread. The south of Buenos Aires province, Argentina, is one of the three regions where hydatidosis is endemic. Although domestic dogs and sheep are considered to be the main hosts for E. granulosus, the potential role of wildlife in the local transmission of E. granulosus has not been investigated. The aim of this study was to estimate the hydatidosis/echinococcosis prevalence in European hare (Lepus europaeus) and Pampas fox (Lycalopex gymnocercus), two abundant species with a strong predator–prey relationship in rural areas of Buenos Aires province using different diagnostic tests. A total of 61 fox intestines were examined, finding that 52 (85.2 %) harbored at least one helminth species. However, no adult or immature form of Echinococcus sp. was found in the intestinal contents. Coproparasitological analysis and Copro–ELISA followed by Copro–PCR were used as supplementary diagnostic tests. Only one (1.7 %) of 59 fecal samples was positive to Taeniidae eggs by coproparasitological analysis, but this same sample was negative by the Copro–ELISA test. The analysis by Copro–ELISA showed 6 of 57 (10.6 %) positive samples, but the Copro–PCR tests carried out on these samples were negative to E. granulosus. A total of 6,808 lungs, 3,576 livers, and 3,542 hearts of hunted hares were examined and palpated, but no structure resembling hydatid cysts were detected. Our results suggest that hares and Pampas foxes are not currently important wild reservoirs of E. granulosus in the studied area.     

Frequency and genetic diversity of Echinococcus granulosus sensu stricto in sheep and cattle from the steppe region of Djelfa,Algeria

Cystic echinococcosis (CE) of humans and animals is caused by various species of Echinococcus granulosus sensu lato. Of these, E. granulosus sensu stricto has the widest geographical distribution and is the most important agent of human cystic echinococcosis. Previous molecular studies showed that E. granulosus s.s. isolates from the Middle East and western Asia exhibit higher intraspecific diversity than those from other parts of the world, which led to hypotheses on the origin of the species in that region. However, various high-endemicity regions have not been sufficiently covered by such studies, including northern Africa as a well-known focus of this parasite. Here, we report data on the mitochondrial cox1 gene (1609bp) sequence diversity of E. granulosus s.s. from Algerian livestock. An abattoir survey of 1278 animals from the Algerian steppe region (Djelfa) resulted in CE prevalence of 13.9% in cattle (n = 266), 5.7% in sheep (n = 975), and 0% in goats (n = 37). All of 125 molecularly examined cyst isolates belonged to E. granulosus s.s. In total, 73 haplotypes were found, only five of which have been previously reported (from the Middle East and Australia). One haplotype sequence (EgAlg01X) was found to contain an insertion of three bases at the end of the gene. To the best of our knowledge, this has not been reported before for Echinococcus spp. Diversity values of our panel of Algerian samples were in the range of those that have been previously reported from the Middle East and far higher than those from elsewhere. This, together with the low number of shared haplotypes, indicates a more complex biogeographical history of this parasite than hitherto assumed.

     

PCR-restriction endonuclease analysis of Mycobacterium avium subsp. paratuberculosis isolates from goats, sheep, and cattle in Jordan

Paratuberculosis is an endemic disease and induces high economical losses in Jordan. There is no information available on genotypic variation of Mycobacterium avium subsp. paratuberculosis (MAP) isolated from animals in Jordan. In this study, we investigated 150 fecal samples from sheep, goats, and cattle for the presence of paratuberculosis using bacterial culture and polymerase chain reaction (PCR)-RFLP analysis of insertion sequence IS1311. Analysis of the results revealed that genotypic information from sheep, goat, and cattle could classify them into cattle or sheep strains. All culture isolates from cattle, 12.5% of the isolates from sheep, and 50% of the isolates from goats were cattle strain, while 87.5% of the isolates from sheep and 50% of the isolates from goats were sheep strain. Sequencing of the IS1311 268?bp PCR product from the three animal species confirmed the different MAP patterns.     

Antibiotic sensitivity of E. coli and Salmonella isolated from different water sources in Kashmir, India

The objective of the present study was to determine the prevalence of Salmonella and Escherichia coli, the serotypes involved and the antibiogram of the various isolates obtained from different sources of water supply (streams, Dal Lake, tube wells and community supply water) in Kashmir, India, during the years 2009–2010. A total of 100 samples, 25 from each source, were taken for the present study. All samples were evaluated for the presence of Salmonella and E. coli, stereotyped and tested for antimicrobial susceptibility. A total of 60 isolates of E. coli and 12 isolates of Salmonella were obtained. The antimicrobial susceptibilities of different isolates of E. coli and Salmonella from different water sources were examined by disc diffusion method. The high in vitro sensitivity was shown by 100 % of E. coli isolates for ciprofloxacin, 95 % for amoxycillin/clavulanic acid, 93.33 % for each of cephotaxime and amikacin and 91.67 % for lavofloxacin and gentamicin. The isolates registered an intermediate response to rifampicin (85.00 %), lomefloxacin (83.33 %) and norfloxacin (83.33 %) and showed resistance to erythromycin (98.33 %). The isolates of Salmonella were 100 % sensitive for gentamicin, 91.67 % for each of cefixime, ciprofloxacin, amikacin and lavofloxacin; 83.33 % for each of cephotaxime and amoxycillin/clavulanic acid and showed resistance to erythromycin (100 %), nalidixic acid (100 %) and rifampicin (91.67 %). The isolates, however, showed an intermediate resistance to lomefloxacin (83.34 %) and norfloxacin (66.67). The high incidence of multi-drug-resistant strains of E. coli and Salmonella is due to injudicious use of antibiotics and exchange of antibiotic resistance among bacterial populations.