Simultaneous quantification of DNA and Ki-67 proliferation-associated antigen was performed using fluorescence image cytometry. In the MCF-7 cell line, the Ki-67 antigen content increases during the cell cycle, and its intranuclear distribution pattern varies. Quantitative evolution of Ki-67 content as a function of nuclear area makes it possible to define several pathways followed by cells going through the 2c compartment. 1) In some cells, the amount of Ki-67 antigen remains constant during G1 (Ki-67 stable pathway), and a characteristic speckled pattern can be observed. 2) In the larger fraction of cells analyzed, there is a postmitotic decrease in the Ki-67 (Ki-67 decrease pathway) content. In this pathway, labeling is located in the nucleoplasm in small nuclei, is located in nucleoli in intermediate-sized nuclei, and is absent from larger nuclei (G0). A progressive increase in Ki-67 content (Ki-67 increase pathway) was observed from intermediate-sized nuclei to S phase nuclei. From these results, we hypothesize that the Ki-67 stable pathway is the G1 phase of newly formed cells going directly to S phase in local optimal conditions of growth and that Ki-67 decrease pathway and Ki-67 increase pathway correspond to cells whose progression to S phase is regulated by extracellular factors.

No abstract

The tropical bont tick, Amblyomma variegatum, is an African tick species which infests livestock and wildlife. It was probably introduced in the central eastern islands of the Caribbean during the 18th or 19th century, with cattle shipped from Senegal. In Africa and the Caribbean, this tick is a vector of heartwater (a rickettsial disease of ruminants) and is associated with acute dermatophilosis (a bacterial skin disease of animals). Until 1948, only Guadeloupe and the neighbouring islands of Marie Galante and Antigua were infested with this tick species. Following increased agricultural commerce between Guadeloupe and Martinique, the latter became infested in 1948. Between 1967 (when the tick was identified in St Croix) and 1988 (when a male tick was reported in St Vincent), fourteen new islands were reached by this tick. Most of the dissemination of the tick to new islands cannot be explained by legal or illegal movements of livestock. Recently-determined circumstantial evidence strongly links the increase in populations of the cattle egret (Bubulcus ibis), a migrating bird established in the Caribbean circa 7960, with increased colonisation of new islands by A. variegatum. Considering the wide range of areas currently occupied by this bird species in the Greater Antilles and on the American mainland, there is a high probability that the tick will also expand its range and invade new areas. Eradication of A. variegatum from the Caribbean and thus from the western hemisphere, and the strengthening of measures to prevent inter-island movements of livestock, would be the only effective means of preventing this threat.

Currently available serological tests for cowdriosis (Cowdria ruminantium infection) in domestic ruminants are hampered by their low specificities because of cross-reactivity with Ehrlichia spp. The use of recombinant major antigenic protein (MAP1) of C. ruminantium for serodiagnosis was investigated. Overlapping fragments of the MAP1 protein were expressed in Escherichia coli and were reacted with sera from sheep infected with either C. ruminantium or Ehrlichia ovina. Two immunogenic regions on the MAP1 protein, designated MAP1-A and MAP1-B, were identified. MAP1-A was reactive with C. ruminantium antisera, E. ovina antisera, and three MAP1-specific monoclonal antibodies, whereas MAP1-B reacted only with C. ruminantium antisera. An indirect enzyme-linked immunosorbent assay (ELISA) based on MAP1-B was further developed and validated with sera from animals experimentally infected with C. ruminantium or several Ehrlichia spp. Antibodies raised in sheep, cattle, and goats against nine isolates of C. ruminantium reacted with MAP1-B. Cross-reactivity with MAP1-B was limited to Ehrlichia canis and Ehrlichia chaffeensis, two rickettsias which do not infect ruminants. Antibodies to Ehrlichia spp. which do infect ruminants (E. bovis, E. ovina, and E. phagocytophila) did not react with MAP1-B. Antibody titers to C. ruminantium in sera from experimentally infected cattle, goats, and sheep were detectable for 50 to 200 days postinfection. Further validation of the recombinant MAP1-B-based ELISA was done with sera obtained from sheep raised in heartwater-free areas in Zimbabwe and from several Caribbean islands. A total of 159 of 169 samples which were considered to be false positive by immunoblotting or indirect ELISA did not react with MAP1-B. In conclusion, recombinant MAP1-B may be a suitable antigen for a sensitive serological test for cowdriosis, with dramatically improved specificity. Cowdriosis (heartwater) is an economically important disease of domestic and wild ruminants caused by the rickettsia Cowdria ruminantium and is transmitted by ticks of the genus Amblyomma (31). The disease is endemic in sub-Saharan Africa and is a major obstacle for upgrading local breeds of livestock with more productive susceptible exotic breeds (31). The disease is also present on at least three Caribbean islands (27) and poses a serious threat to livestock production on the North and South American mainlands (2). Serological tests that have been developed for cowdriosis are based on detection of antibodies by immunofluorescence (11, 21, 28) or by indirect enzyme-linked immunosorbent assay (iELISA) with in vitro-cultured C. ruminantium organisms (24). However, cross-reactivity between C. ruminantium antigens and antibodies to several Ehrlichia spp. has been observed (9, 11, 15, 23). In addition, a competitive ELISA (cELISA) (19) and an immunoblotting technique (23) have been developed. These tests are based on an immunodominant 32-kDa surface protein of C. ruminantium designated major antigenic protein (MAP1 [1]; it was formerly known...